Interesting side note....
http://www.ibiblio.org/herbmed/eclectic/kings/acidum-gall.html - the "elder Robiquet"?
posted by Ari at 5:30 PM
Thursday, September 29, 2005
Wednesday, September 28, 2005
Translation of Vauquelin Article
Discovery of a novel fundamental vegetable substance in Asparagus (Sativus, Linnean)
Upon examining more precisely the products of plant matter which have formerly not been done, modern chemists have distinguished a large number of species (types) unknown by ancient people. But for a long time, I believe, we have not found in the vegetables an immediate substance also as singularly interesting as that which we discuss here.
During the past year(summer), M. Robiquet, a young chemist who combines the solidity of reasoning with a great skill in the experiments submitted, at the invitation of M. Parmentier, has done chemical analysis on the juice of Asparagus whose interesting results he has submitted to the Annals of Chemistry.
Having given up (having been abandoned) in my laboratory, during a voyage that it (he) made, a certain quantity of asparagus juice concentrated by evaporization, I had observed a somewhat large number of crystals, among which two apparently new substances appeared, which had a form, a transparence, and a different smell/flavor that made it easy for me to separate them.
One of these chemical species - perfectly white and trasnparent when it had been recrystallized several times - had a slightly nauseating sharp smell/flavor which caused the mouth to water; it ('s crystals) are durable, brittle and of a regular form.
The other chemical species - equally white, was not also transparent nor durable nor recrystallized in the same form; it was on the contrary without consistancy, crystallized in fine needles, had a noticably sweet essense analagous to a (botanical) manna (true essense of asparagus?).
M. Robiquet, in doing the work previously mentioned, had perceived the first of these substances but he believed that is was an ammonium salt because he had not then been able to procure but a very small quantity and imperfect purification, it retained according to all appearance, between its blades some traces of ammonia base-salt to which the asparagus juice bound which misled him.
Since this time, we have together submitted this substance to new experiments, which yielded the prinicple chemical species. The form that it takes on in crystallization, after M. Hauy, to whom we have given a certain quantity, derived of it a right rhoboidal prism in which the large angle of the base is approximately 130 degrees. The edges of this base and the two angles situated at the extremity of its large diagonal are replaced by facets.
This substance is moderately soluble in water and its dissolution does not give any acidity nor basicity: the infusion of gallnuts (gallic acid), lead acetate, ammonium oxalate, barium chloride (?!?), and potassium hydrogen sulfate (?) did not change to the (degree of) dissolution of this substance. It was not soluble in alcohol.
These tests indicated that the matter in which it acts is not an alkaline earth salt. We pulverized a sample with caustic potassium and a small amount of water for seeing if it would neutralize ammonia, but we did not perceive any discernable trace: the postassium (yielded?) appeared to us more soluble in water.
This indicates thus that it contained neither (earth) [historically, essential elements were considered to consist of "earth" among the "elements" - Air, Fire, Water, and in some cases oil and salt were also "elements.] (?) nor ammonia - we have investigated the existance of alkalis, and for that we have done burn tests on a somewhat large quatity in a platinum crucible , it was initially considerably puffy (effervescent?) in exhaling piquant vapors which affected the eyes and the nostrils like wood smoke. It yielded a great deal of charcoal that did not have a threshold of (smell?), and which did not leave after its incineration but an almost imperceptable trace of earth (element), which is certainly strange(novel?).
At the end of the composition of this material, the odor which emanates is analagous to that of organic matter and also a little like ammonia.
Nitric acid decomposes this substance, it emits nitrous gas, the mother liquor takes a yellow color, a bitter flavor like organic substances. When the action of nitric acid is achieved, the lime breaks down abundantly of the ammonia of the mother liquor.
This alkali is thus formed in the operation which we have just described since the substance from asparagus does not show any characteristics detectable before.
This substance is not an acid since it does not redden litmus (tincture) and because it does not have the essense infected to all these bodies, marked in a positive or negative degree.
It is not a neutral salt because it doesn not contain earth or salt (elements), but as it yields by means of fire ther same products as vegetables, we are obliged to view it as a essential constituent of asparagus.
It is probable that it is composed likethose substances of hydrogen, oxygen, and carbon in the particular proportions it is no less probable that it also has a small constituent of nitrogen; it is less evident indicated by the odor that it decomposes by heat and the ammonia that it forms with nitric acid.
Althought we have obtained a somewhat large quantity of this substance, we have not been able to submit it to a very large number of tests because most of our sample is misplaced in our lab. and it does not rest with us that the smalle portion that we have given to M. Hauy, for determining the crystal form: nevertheless we believed we had to share part with the Institut in order to take date, we propose to continue our work at once when the asparagus arrives (harvests?).
We will also investigate if this singular substance does or does not exist in other vegetables.
As for the sweet material that we have also isloated, in the asparagus juice, we do not sufficiently know to which category of sugar it most resembles - we believe it depends what it is to the exudate.
One can thus say for certain, that in addition to the essential substances discovered in asparagus juice by M. Robiquet, there exists a substance that is crystallizable like salts, and which is certainly not an acid, nor a neutral salt, and in which solution in water is not affected by any of the reactions which are ordinarily used to recognize the presence and nature of salts dissolved in water, and another sweet substance which would seem to be analagous to manna (exudate?).
Upon examining more precisely the products of plant matter which have formerly not been done, modern chemists have distinguished a large number of species (types) unknown by ancient people. But for a long time, I believe, we have not found in the vegetables an immediate substance also as singularly interesting as that which we discuss here.
During the past year(summer), M. Robiquet, a young chemist who combines the solidity of reasoning with a great skill in the experiments submitted, at the invitation of M. Parmentier, has done chemical analysis on the juice of Asparagus whose interesting results he has submitted to the Annals of Chemistry.
Having given up (having been abandoned) in my laboratory, during a voyage that it (he) made, a certain quantity of asparagus juice concentrated by evaporization, I had observed a somewhat large number of crystals, among which two apparently new substances appeared, which had a form, a transparence, and a different smell/flavor that made it easy for me to separate them.
One of these chemical species - perfectly white and trasnparent when it had been recrystallized several times - had a slightly nauseating sharp smell/flavor which caused the mouth to water; it ('s crystals) are durable, brittle and of a regular form.
The other chemical species - equally white, was not also transparent nor durable nor recrystallized in the same form; it was on the contrary without consistancy, crystallized in fine needles, had a noticably sweet essense analagous to a (botanical) manna (true essense of asparagus?).
M. Robiquet, in doing the work previously mentioned, had perceived the first of these substances but he believed that is was an ammonium salt because he had not then been able to procure but a very small quantity and imperfect purification, it retained according to all appearance, between its blades some traces of ammonia base-salt to which the asparagus juice bound which misled him.
Since this time, we have together submitted this substance to new experiments, which yielded the prinicple chemical species. The form that it takes on in crystallization, after M. Hauy, to whom we have given a certain quantity, derived of it a right rhoboidal prism in which the large angle of the base is approximately 130 degrees. The edges of this base and the two angles situated at the extremity of its large diagonal are replaced by facets.
This substance is moderately soluble in water and its dissolution does not give any acidity nor basicity: the infusion of gallnuts (gallic acid), lead acetate, ammonium oxalate, barium chloride (?!?), and potassium hydrogen sulfate (?) did not change to the (degree of) dissolution of this substance. It was not soluble in alcohol.
These tests indicated that the matter in which it acts is not an alkaline earth salt. We pulverized a sample with caustic potassium and a small amount of water for seeing if it would neutralize ammonia, but we did not perceive any discernable trace: the postassium (yielded?) appeared to us more soluble in water.
This indicates thus that it contained neither (earth) [historically, essential elements were considered to consist of "earth" among the "elements" - Air, Fire, Water, and in some cases oil and salt were also "elements.] (?) nor ammonia - we have investigated the existance of alkalis, and for that we have done burn tests on a somewhat large quatity in a platinum crucible , it was initially considerably puffy (effervescent?) in exhaling piquant vapors which affected the eyes and the nostrils like wood smoke. It yielded a great deal of charcoal that did not have a threshold of (smell?), and which did not leave after its incineration but an almost imperceptable trace of earth (element), which is certainly strange(novel?).
At the end of the composition of this material, the odor which emanates is analagous to that of organic matter and also a little like ammonia.
Nitric acid decomposes this substance, it emits nitrous gas, the mother liquor takes a yellow color, a bitter flavor like organic substances. When the action of nitric acid is achieved, the lime breaks down abundantly of the ammonia of the mother liquor.
This alkali is thus formed in the operation which we have just described since the substance from asparagus does not show any characteristics detectable before.
This substance is not an acid since it does not redden litmus (tincture) and because it does not have the essense infected to all these bodies, marked in a positive or negative degree.
It is not a neutral salt because it doesn not contain earth or salt (elements), but as it yields by means of fire ther same products as vegetables, we are obliged to view it as a essential constituent of asparagus.
It is probable that it is composed likethose substances of hydrogen, oxygen, and carbon in the particular proportions it is no less probable that it also has a small constituent of nitrogen; it is less evident indicated by the odor that it decomposes by heat and the ammonia that it forms with nitric acid.
Althought we have obtained a somewhat large quantity of this substance, we have not been able to submit it to a very large number of tests because most of our sample is misplaced in our lab. and it does not rest with us that the smalle portion that we have given to M. Hauy, for determining the crystal form: nevertheless we believed we had to share part with the Institut in order to take date, we propose to continue our work at once when the asparagus arrives (harvests?).
We will also investigate if this singular substance does or does not exist in other vegetables.
As for the sweet material that we have also isloated, in the asparagus juice, we do not sufficiently know to which category of sugar it most resembles - we believe it depends what it is to the exudate.
One can thus say for certain, that in addition to the essential substances discovered in asparagus juice by M. Robiquet, there exists a substance that is crystallizable like salts, and which is certainly not an acid, nor a neutral salt, and in which solution in water is not affected by any of the reactions which are ordinarily used to recognize the presence and nature of salts dissolved in water, and another sweet substance which would seem to be analagous to manna (exudate?).
posted by Ari at 1:01 PM
Monday, September 26, 2005
Nutrition of Amino Acids
William C. Rose (UIUC and later UC Berkeley?) made a lot of discoveries pertaining to the nuritive effects of AA's or lack of them. Was the first? to describe which AA's were and were not essential. Working on getting a comp. list of his papers - have a number saved on office PC.
The dog nutrion paper (Science, 1939) describes essential AA's and points to earlier papers.
The arginine deficeincy paper (Jrnl Biological Chemistry, 1930) describes even earlier research.
The dog nutrion paper (Science, 1939) describes essential AA's and points to earlier papers.
The arginine deficeincy paper (Jrnl Biological Chemistry, 1930) describes even earlier research.
posted by Ari at 4:16 PM
Peptide Bond - Description
First described in terms of bond lengths and general shape by Linue Pauling 1948-1952. Looking for resources on Pauling.
posted by Ari at 4:15 PM
Amino Acids - Peptide Bond Discovery
Supposedly - Emil Fischer first described the peptide bond between 1899 and 1901. According to internet sources, he worked with Fourneau on this. Looking for papers from Fischer at that time.
posted by Ari at 4:13 PM
Wednesday, July 20, 2005
Test 2 Q - Answers
1. Actin filaments form (like "tinkertoys") from actin monomers - assesmbling on their assembly (+) end (torwards the cell membrane) and where ATP cap is in equilibrium filament and disassembling as needed on their disassembly (-) end (ADP cap end in equilibrium). Don't come apart because the membrane has an excess of lipids that have lateral motion around the membrane. When actin pushes out aganist the membrane, the membrane phospholipids can pull apart from one another somewhat (although not apart enough to overcome the Van der Waals and hydrophobic attractions that keep the membran intact) forming spaces between lipids that the excess lipids from other areas of the membrane can move into.
2. Compare and contrast Actin, Microtubules, and Intermediate Tubules. Be complete - use all the information I told you in class.
Actin:
Structure / Polarity - "donut" shaped alpha, beta, gamma monomers 42 kD, ~375 AA's differ from one another by 4 -5 AA's (?!?!), each has a pocket containing ATP/ADP. Polarity of each mono. gives filament polarity. ATP and ADP caps ends, 6 nm in diameter filaments.
Assembly / Disassembly - ATP cap is ass end, ADP cap w/ less binding NRG is diass end. At eq. 1 mono added for each taked - gives treadmilling in vitro and vivo
In vitro conditions - will ass / diass in vitro and in vivo. Know what's needed for this...
Nucleation cites - actin forms at nuc. sites at Memb Anchor proteins just inside PM.
Intracellular prefs - cell. conditions - treadmillling occurs as actin filaments are assembled as "tinkerotys" to extend cell fila. and lamelli. don't bk PM b/c extra lipids in memb. come to fill spaces when actin pushes out
Assoc. Proteins - Myosin I, Myosin II - light and heavy chains (draw a ceduceus) 220 kD each heavy, 22 kD each light chain. Bundled to form "bouquets" of fibers layered between bands of actin, and are Ca 2+ dependent activated to contract muscles. (Chk this). Villin - first thought to be actin bunding protein in brush barrier cells, came out at 72 kD on SDS-PAGE. BUT Villin works based on [] of free Ca 2+. Fimbrin at 68 kD turned out to be the bundling protein. Capping proteins (?!?!?!) - let cell control start and stop of creation of actin filaments (?!?!?).
MTs:
Structure / Polarity - each TUBULIN monomer is actually a homodimer? with alpha and beta subunits, each 55 kD and ~ 450 AA's in length. These polymerize together in a "braided" chain - end to end the chain of monomers forms a protofilament. as they braid around to form a MT - they form 13 alternating subunits around an open space (draw better) than is 25 nm on its outer cirum and 15 nm on the inner, mono. ring is ~ 5 nm in width. The units each pocket a GTP/GDP - only the beta subunit transforms to GDP. Monomers and hence the filament have polarity.
Assembly / Disassembly - have + and - ends, disintigrate rapidly in cell.
In vitro conditions - MTs treadmill in vitro but NOT in the cell. - extracted from cow brains, What are conditions in vitro - Know....
Nucleation cites - Centrosome - don't touch the centrioles, but will not form witout centriols in animal cells??!?!? Have a "cap" ?!?!?! inside the centrosome- this is the (-) end. (+) ass. end out into cytoplasm from centrosome center.
Intracellular prefs - do NOT treadmill in cell - disintegrate too rapidly. Grow out to near PM. Act as "roads to Rome" of the cell.
Assoc. Proteins - Dynein connects like legs outside of vesicles / organelles to walk along MT to (-) Kinesin - same, but out to (+) part of MTs.
IF's:
Structure / Polarity - Monomeric units consist of 4 strands of peptides that alternate btw 5 linear and 4 alpha helical regions. Hydophobic areas (at the linear parts?) cause the strands to bond more tightly than almost another natural substance in animal cells. b/c of alternating "lines of fracture" - IF's are "brick shithouses" of natural fibers. Each of 4 strands is polar with N and C term ends. Brought together in 4 they "lose" polarity, however.
Assembly / Disassembly - 4 strands come together w/ hydrophobic regions, DO NOT disass.
In vitro conditions - have to boil at 100 deg C in 2% SDS and 4 M Urea to denature.
Nucleation cites - right at Nuclear membrane - stands extrude through pores to outside and bond immeidately to outside of nuclear memb. Transported back in thru pores to form Nuc. lamins. This forms with "stainless steel jockstrap" springy cage that protects the Nuc. from crushing.
Intracellular prefs - 4 strand monomers twist together to form long strands that then bundle in 4 cables which circle around in 8 bundles to form the IF.
Assoc. Proteins - we didn't cover yet? Not used to transport - no polarity!!!!!
3. ATP, GTP, CTP, TTP. ATP is found in actin, GTP in microtubules. It is not known exactly why actin filaments use ATP in particular for a non-NRG generating purpose, but ATP gets converted into ADP by an enzyme. These conversions, happening randomly, eventually form the ATP and ADP caps - the ATP is the "assembly end", and ADP the disassembly end (the ADP has 1 less negative charge and dissociates more readily).
In MT's, in the Tubulin monomer (actually a heterodimer), both the (joined) alpha and beta subunits start off containing GTP, but only the beta subunit GTP gets converted to GDP. More needed ...
3. What are some examples of nucleotide tri-phosphates? Where specifically to we find the ones we talked about in class? How are they used?
ATP, GTP CTP, TTP. ATP is used in actin filaments, not as NRG source, but to mark the filament for ass / disass ends. GTP is found in tubulin monomers of MT's. It also signifies polarity of the MT.
4. What is polarity when in biological macromolecules? Why is it dissimilar to electrical charge? What causes polarity?
As in class - polarity means different directionality. It is not electrical charge b/c doen't connote free electrons moving in flow from one place to another, but is the position change of monomers as the cycle through an actin filiment. Or the directionality that vesicles are "walked" along a cytoskelatal fiber to move throughout a cell. This only happens in one direction for a given molecular motor. Polarity or lack of it arise from the polarity of monomers.
5. What are some of the "families" seen in cellular matix systems? What are some of the members? How are the families organized?
Actin filaments have alpha, beta, gamma types, but only on family member makes up a single actin filament (?!?!?). In contrast, in tubulin, alpha and beta units are attached together to make a monomer - so there is intermingling of familial types.
6. What makes IF's the "brick shithouses" of the cell biology world?
The monomers align in a staggered conformation to form a filament, much like brick walls are staggered. This non-grid alignment means that there are no single potential lines of fracture directly across or along the filament, making IF's even more strong. Additionally, there are hydroprobic strips every 7 repeats (?!?!?!) that make the assembled IF extermely strong.
7. How can we verify the polarity of MT's in the cell?
In vitro, can use free monomers which under cell cond. will form protofilament "wings" off the MT. Through thin-slice EM ALONG the MT, these were also seen. Turns out CW is to (-) end - to the centrosome. CCW is to (+) end - to PM.
8. How did scientists first test how myosin works in cells? What steps did they do to test this?
2. Compare and contrast Actin, Microtubules, and Intermediate Tubules. Be complete - use all the information I told you in class.
Actin:
Structure / Polarity - "donut" shaped alpha, beta, gamma monomers 42 kD, ~375 AA's differ from one another by 4 -5 AA's (?!?!), each has a pocket containing ATP/ADP. Polarity of each mono. gives filament polarity. ATP and ADP caps ends, 6 nm in diameter filaments.
Assembly / Disassembly - ATP cap is ass end, ADP cap w/ less binding NRG is diass end. At eq. 1 mono added for each taked - gives treadmilling in vitro and vivo
In vitro conditions - will ass / diass in vitro and in vivo. Know what's needed for this...
Nucleation cites - actin forms at nuc. sites at Memb Anchor proteins just inside PM.
Intracellular prefs - cell. conditions - treadmillling occurs as actin filaments are assembled as "tinkerotys" to extend cell fila. and lamelli. don't bk PM b/c extra lipids in memb. come to fill spaces when actin pushes out
Assoc. Proteins - Myosin I, Myosin II - light and heavy chains (draw a ceduceus) 220 kD each heavy, 22 kD each light chain. Bundled to form "bouquets" of fibers layered between bands of actin, and are Ca 2+ dependent activated to contract muscles. (Chk this). Villin - first thought to be actin bunding protein in brush barrier cells, came out at 72 kD on SDS-PAGE. BUT Villin works based on [] of free Ca 2+. Fimbrin at 68 kD turned out to be the bundling protein. Capping proteins (?!?!?!) - let cell control start and stop of creation of actin filaments (?!?!?).
MTs:
Structure / Polarity - each TUBULIN monomer is actually a homodimer? with alpha and beta subunits, each 55 kD and ~ 450 AA's in length. These polymerize together in a "braided" chain - end to end the chain of monomers forms a protofilament. as they braid around to form a MT - they form 13 alternating subunits around an open space (draw better) than is 25 nm on its outer cirum and 15 nm on the inner, mono. ring is ~ 5 nm in width. The units each pocket a GTP/GDP - only the beta subunit transforms to GDP. Monomers and hence the filament have polarity.
Assembly / Disassembly - have + and - ends, disintigrate rapidly in cell.
In vitro conditions - MTs treadmill in vitro but NOT in the cell. - extracted from cow brains, What are conditions in vitro - Know....
Nucleation cites - Centrosome - don't touch the centrioles, but will not form witout centriols in animal cells??!?!? Have a "cap" ?!?!?! inside the centrosome- this is the (-) end. (+) ass. end out into cytoplasm from centrosome center.
Intracellular prefs - do NOT treadmill in cell - disintegrate too rapidly. Grow out to near PM. Act as "roads to Rome" of the cell.
Assoc. Proteins - Dynein connects like legs outside of vesicles / organelles to walk along MT to (-) Kinesin - same, but out to (+) part of MTs.
IF's:
Structure / Polarity - Monomeric units consist of 4 strands of peptides that alternate btw 5 linear and 4 alpha helical regions. Hydophobic areas (at the linear parts?) cause the strands to bond more tightly than almost another natural substance in animal cells. b/c of alternating "lines of fracture" - IF's are "brick shithouses" of natural fibers. Each of 4 strands is polar with N and C term ends. Brought together in 4 they "lose" polarity, however.
Assembly / Disassembly - 4 strands come together w/ hydrophobic regions, DO NOT disass.
In vitro conditions - have to boil at 100 deg C in 2% SDS and 4 M Urea to denature.
Nucleation cites - right at Nuclear membrane - stands extrude through pores to outside and bond immeidately to outside of nuclear memb. Transported back in thru pores to form Nuc. lamins. This forms with "stainless steel jockstrap" springy cage that protects the Nuc. from crushing.
Intracellular prefs - 4 strand monomers twist together to form long strands that then bundle in 4 cables which circle around in 8 bundles to form the IF.
Assoc. Proteins - we didn't cover yet? Not used to transport - no polarity!!!!!
3. ATP, GTP, CTP, TTP. ATP is found in actin, GTP in microtubules. It is not known exactly why actin filaments use ATP in particular for a non-NRG generating purpose, but ATP gets converted into ADP by an enzyme. These conversions, happening randomly, eventually form the ATP and ADP caps - the ATP is the "assembly end", and ADP the disassembly end (the ADP has 1 less negative charge and dissociates more readily).
In MT's, in the Tubulin monomer (actually a heterodimer), both the (joined) alpha and beta subunits start off containing GTP, but only the beta subunit GTP gets converted to GDP. More needed ...
3. What are some examples of nucleotide tri-phosphates? Where specifically to we find the ones we talked about in class? How are they used?
ATP, GTP CTP, TTP. ATP is used in actin filaments, not as NRG source, but to mark the filament for ass / disass ends. GTP is found in tubulin monomers of MT's. It also signifies polarity of the MT.
4. What is polarity when in biological macromolecules? Why is it dissimilar to electrical charge? What causes polarity?
As in class - polarity means different directionality. It is not electrical charge b/c doen't connote free electrons moving in flow from one place to another, but is the position change of monomers as the cycle through an actin filiment. Or the directionality that vesicles are "walked" along a cytoskelatal fiber to move throughout a cell. This only happens in one direction for a given molecular motor. Polarity or lack of it arise from the polarity of monomers.
5. What are some of the "families" seen in cellular matix systems? What are some of the members? How are the families organized?
Actin filaments have alpha, beta, gamma types, but only on family member makes up a single actin filament (?!?!?). In contrast, in tubulin, alpha and beta units are attached together to make a monomer - so there is intermingling of familial types.
6. What makes IF's the "brick shithouses" of the cell biology world?
The monomers align in a staggered conformation to form a filament, much like brick walls are staggered. This non-grid alignment means that there are no single potential lines of fracture directly across or along the filament, making IF's even more strong. Additionally, there are hydroprobic strips every 7 repeats (?!?!?!) that make the assembled IF extermely strong.
7. How can we verify the polarity of MT's in the cell?
In vitro, can use free monomers which under cell cond. will form protofilament "wings" off the MT. Through thin-slice EM ALONG the MT, these were also seen. Turns out CW is to (-) end - to the centrosome. CCW is to (+) end - to PM.
8. How did scientists first test how myosin works in cells? What steps did they do to test this?
posted by Ari at 3:39 PM
Test 2 Possible Q's
1. By what mechanism does the cell extend filapodia and lamellipodia? Why don't membranes come apart when this happens?
2. Compare and contrast Actin, Microtubules, and Intermediate Tubules. Be complete - use all the information I told you in class.
3. What are some examples of nucleotide tri-phosphates? Where specifically to we find the ones we talked about in class? How are they used?
4. What is polarity when in biological macromolecules? Why is it dissimilar to electrical charge? What causes polarity or lack therof?
5. What are some of the "families" seen in cellular matix systems? What are some of the members? How are the families organized?
6. What makes IF's the "brick shithouses" of the cell biology world?
7. How can we verify the polarity of MT's in the cell?
8. How did scientists first test how myosin works in cells? What steps did they do to test this?
9.
2. Compare and contrast Actin, Microtubules, and Intermediate Tubules. Be complete - use all the information I told you in class.
3. What are some examples of nucleotide tri-phosphates? Where specifically to we find the ones we talked about in class? How are they used?
4. What is polarity when in biological macromolecules? Why is it dissimilar to electrical charge? What causes polarity or lack therof?
5. What are some of the "families" seen in cellular matix systems? What are some of the members? How are the families organized?
6. What makes IF's the "brick shithouses" of the cell biology world?
7. How can we verify the polarity of MT's in the cell?
8. How did scientists first test how myosin works in cells? What steps did they do to test this?
9.
posted by Ari at 3:21 PM
Tuesday, July 12, 2005
Lecture 1 Notes 7/5
Possible Q's
What are the origins of cell biology? Was there agreement among all scientists in early Cell Biology? If not, what were the differences? What are significant characteristics?
What am I saying when I say there are limits to technology? What are some examples of this? How are some ways we work around these?
What types of "technology" do cells employ? What do these technologies tell us about cellular evolution?
Tell me something about dimensionality? What does this have to do with cells?
Define PHENOMENOLOGY. What does it have to do with the study of cells?
3) Why was is lucky for cell culture studies that many of the early efforts at cell cultures were attempted with cells from rodents such as mice?
If I said: Because rodents cells grow readily in culture and much more easily when little was know about cell cultures. Also they become cancerous at a higher rate than the cells of larger mammals. IS THIS ENOUGH - OR DO I NEED TO BREAK DOWN 'CANCEROUS' into "IMMORTAL" and "TRANSFORMED"?
4) What survives better in cell culture systems, cells from adults or cells from early developmental stages such as embryos? Explain your answer.
5) Starting with a chunk of tissue removed from an organism, explain how a cell line is obtained. In this answer you should list a complete, detailed procedure including all the elements discussed in class.
6) Concering your answer to number 5 - what would happen if instead of using serum to provide growth factors to the culture medium I instead added the exact growth factors to promote growth of the cells I specifically wanted to obtain. NOTE: be cautions in your answer because this question is a bit tricky.
Assume you have a suspension culture of cells growing from a tumore. You start the culture with 10 cells in it and the suspension culture has 100 mLs of medium in it. (In case you don't remember a suspension culture is s culture where the cells are gently mixed so that they are floating around.The currents in the mdeium prevent them from settling down onto the bottom of the culture plate). Answer the following questions below (9-12) with regard to this experiement.
9) How many cells will be present in the plate 3 days later? WHAT"S THE ANSWER? DID WE EVER GET A RATE OF GROWTH? CANCER CELLS DIVIDE in 18 HRS. IS THAT IT?
4 x 18 = 72 hrs. 10 > 20 > 40 > = 80 ?
10) If you wanted to isolate 1 cell out of the suspension culture to start a new suspension culture from a single cell, describe two methods discussed in class to perform this task.
11) What would you predict would happen if you took the initial suspension culture and turned off the mechanism that gently mixed the cells?
12) How would you test your prediction?
13) Assume that you have a cell walking on a substrate. With regard to that cell, describe the various parts of the cells and how these structures give you information about the directionality of the cell. IF I DREW A PICTURE OF THE NON-WALKING CELL TO SHOW NO MOVEMENT - WOULD THIS BE INCORRECT? Or INCORRECT WITHOUT?
14) Describe the process that has to occur in a normal cell if it is to become both anchorage-independent for growth and immotal? I suggest you use a diagram to answer this question.
-----------------------
Outline:
- Intro
- Origin of Cell Biology
- Biomolecular mechanisms / machines
- Problems with Dimenstionality
- Behavior of Cells
- Cell Walking - Culture + in Biology
- Cells as organisms
- How do you look at cells?
------------------------------
Cell Biology
-----------------
It can be seen through 2 historical perspectives
1) Biochemical perspective:
Biochemists were one group of researches who began CB. They homogenized cells first and tested these parts of cells
2) Morphologists:
Used ELETRON MICROSCOPY - they fixed and viewed cells under a microscope to find out more about them.
In the 1950's CB had groups involved like the (ASM) Microbiologists - wanted to understand "how cells work"
--They were trying to understand what cells DO. Cells were NOT intuitive - scientists didn't know how they worked
"Evolution designed cells" - if cells had desirable characteristics, they lived. If they didn't, they died
What are "Biomolecular machines" and what do they do? How do we understand biomolecular machines?
First approach is DESCRIPTIVE:
If we want to describe anything, what do we do? We look at it, look at its structures. Macroscopic observation means looking at something overall and describing what we know empirically by observing with our eyes, senses.
BUT - this doesn't work with CELLS. Why not? Not because:
- Cells are small, TINY - we can't actually see them - so we have to use MACHINES to do this.
- Electron Microscopy is a technology we use - BUT - our view are only FREEZE FRAME picutres
- It took 40+ years to figure out some basic structures.
SO - Cells are THEORETICAL because of out limits of observation
Given all this, since we have to have indirect approaches to study cells - theoretical ways - scientists came up with approaches to do so.
Cliques arose between the BIOCHEMISTS and MICROBIOLOGISTS camps and the didn't agree upon which approach was best - do we study the minute details - the individual pieces because we couldn't look at the parts in an intact cells, or did we look without a lot of detail at whole cells?
- Example: Microfilaments, a basic structure in cells - was huge source of contention. I remember going to a conference where grown men called each other names and used four-letter words over whether microfilaments exisited. MIC's insisted that they could look right at cells and see them. BCHs laughed and said "you're crazy"- when we break the cells up, there is absolutely no evidence they exist.
Biochemists like to isolate single, pure parts of a cells or molecules and study, test these. They take a tube with a receptacle and a plunger, and plunge, plunge to break cells apart and then centifuge. At the bottom of the tube you get "gemisch" and plasma above that. So they take these cells parts from the bottom and study them. What might be a limitation for this? It's one approach, BUT - "all cells come from cells". When you do this - are you studying a live cell any more? Can you tell exactly what it's doing? Can you tell by observing - what the cell is doing? NO - it's DEAD!
REVIEW THIS - CAN YOU CLARIFY WHAT YOU MEANT BY THE LIGHT SWITCH and MARKERS?
So - to see a live, or at least entatct, cell, you put it under a Microscope, you look at it. What do you see [refers to a drawing on board] let's say we have these structures and parts. What is "uniform" vs. "non-uniform" - what are separate distinct stuctures? What do they do? We cam to realize over time that these were separate "machines" with specific functions.
Example: let's say you come into a room you've never been in before. You see the light switch. Even if you've never seen one like this before, could you guess at what is does? Could you guess that it at least does something specific because it is discrete and it's there? You walked into the room, and you turned it on. It is distinct and has a function. You did these in sequence. The same with these board markers.
This is the same thing with cells. We look at cells and we can tell that because there are structures, organization, distinct or discrete characteristics, that these parts and stuctures have function. We may not know what they do - we still don't know what some of these things do - but we know that the cells have evolved to have them as "machines".
This is the same as biotechnology - they take these machines, the mechanisms of cells and do tasks - like the light switch - ON DEMAND and OUT OF SEQUENCE, to meet the needs the researchers are looking for.
DIMENSIONALITY
-----------------------------
WHICH EM IS THE BEST? SEM? TEM?
- Cells are SMALL
- Our understanding of cells is ABSTRACT
- ELECTRON MICROSCOPY is in STILL FRAMES (DEAD CELLS)
- Thin 2-D slices is what we see in Trasnmission Electron Microscopy. So it would be like you looked at me through thin slices - would you be able to get a good idea of how I looked if this is what you saw?
- Cytologic FIXATION KILLS Cell
- Scanning Electon Microscopy is another method that was developed and it addressed some of the drawbacks of TEM - it shows 3-D views of cells at the surface but at one time was not as high resolution as now. SEM is better, higher resolution than LIGHT MICROSCOPY - you can see filaments such as FILIPODIA which are only 0.2 x 10 micrometers - much smaller structures than can be seen with LM. So - SEM has about a 0.2 micrometer resolution.
Drawbacks to SEM:
- Can't see within cells
- Can't see motion
LIGHT MICROSCOPY (LIVING CELLS):
- Bright field - use stained slides
- Phase constast
- Differential Interferometry?!?! Contrast
---------------------------------------------------------------
CELL BEHAVIOR - phenomenological behavior, involves how cells are predicted to act, based on what we've observed of their behavior. For example, if I see a cell moving and it stops, I can predict with good accuracy what the cell will do next.
So - if we drop cells into a culture, and they are suspended and free to move about - what shape will they assume?
- SPHERE!!! Why is this? Because the sphere is a shape has the smallest sruface area with the largest volume. All cells in culture assuma SPHERICAL shape and fall to the bottom of their container. What's interesting is that when they hit that bottom, they being to explore. Let me explain....
First, though - tell me, what are some examples of cell types? [From stus: hepatocytes, hempcytes. These react DIFFERENTLY to their enrivonment. For example - if I put a rabbit into a box - what happens? It will go to the middle of the box. If I do the same with a rat - what do you predict will happen? It will run to the corner and cower. These are different species, but just by knowing what they are and having observed them, I can predict with great certainty what they will do]
So - we have cells, say our heptocytes that hit the botton of the container - what do they do? They are sphereical as they fall - but look at the picture [Has a Sphere, half-sphere, half-flat, and somewhat flat cells]
They touch the bottom, and:
FILIPODIA: - the cell initially extends these in all directions. They are tiny hair-like, like fingers to "feel" their environment. 0.2 micometers in diameter, 2-10 micrometers in length. So, the cell extends these out to feel. As it does this, the cell begins to touch the bottom of the container. The cells does this for a while and then stops 2-3 minutes - a pause. Then...
LAMELLIPODIA: once the cell senses the surface with FILIPODIA and pauses, it extends a larger stucture like a leg, engulfing "swallowing" the FILIPODIA as it extends. As the cell extends a LAMELLIPODIA, is secretes a substances onto the surface of the container - the "cellular equivalent of superglue" so that it sticks and continues extending until FLAT and like a pancake. It may stay there, or it may begin to "WALK". [Note the drawings of a "pancake" with a hump in the middle - NONW WALKING and one with the hump to a side - WALKING.] The lamellipodia then assembles actin and miosin filaments together to form the "cellular equivalent of a muscle" which contracts to MOVE THE CELL?!?!? then sends another lamellipodia out in the direction it wants to go. It will only do this in the direction of the surface of the container. The cell won't waste ATP trying to extend lamellipodia in a diection it won't be able to move in.
As the cell contracts and WALKS, it leaves behind RETRACTION FIBERS - which are the superglue form where the lamellipodia attached and small blebs of plasma membrane that trail behind. Eventually these release and merge (mostly) back into the cell's membrane and cytoplasm. After the retraction fiber releases, there will still be traces of superglue and membrane left - MICROEXUDATES.
As the cell moves, it "drags" the Nucleus (which causes the bulge) using ATP ONLY in the lamellipodia. (The cell doesn't waste ATP by forming and contracting "muscles" in the part of the cell around the trailing Nucleus.)
What are the origins of cell biology? Was there agreement among all scientists in early Cell Biology? If not, what were the differences? What are significant characteristics?
What am I saying when I say there are limits to technology? What are some examples of this? How are some ways we work around these?
What types of "technology" do cells employ? What do these technologies tell us about cellular evolution?
Tell me something about dimensionality? What does this have to do with cells?
Define PHENOMENOLOGY. What does it have to do with the study of cells?
3) Why was is lucky for cell culture studies that many of the early efforts at cell cultures were attempted with cells from rodents such as mice?
If I said: Because rodents cells grow readily in culture and much more easily when little was know about cell cultures. Also they become cancerous at a higher rate than the cells of larger mammals. IS THIS ENOUGH - OR DO I NEED TO BREAK DOWN 'CANCEROUS' into "IMMORTAL" and "TRANSFORMED"?
4) What survives better in cell culture systems, cells from adults or cells from early developmental stages such as embryos? Explain your answer.
5) Starting with a chunk of tissue removed from an organism, explain how a cell line is obtained. In this answer you should list a complete, detailed procedure including all the elements discussed in class.
6) Concering your answer to number 5 - what would happen if instead of using serum to provide growth factors to the culture medium I instead added the exact growth factors to promote growth of the cells I specifically wanted to obtain. NOTE: be cautions in your answer because this question is a bit tricky.
Assume you have a suspension culture of cells growing from a tumore. You start the culture with 10 cells in it and the suspension culture has 100 mLs of medium in it. (In case you don't remember a suspension culture is s culture where the cells are gently mixed so that they are floating around.The currents in the mdeium prevent them from settling down onto the bottom of the culture plate). Answer the following questions below (9-12) with regard to this experiement.
9) How many cells will be present in the plate 3 days later? WHAT"S THE ANSWER? DID WE EVER GET A RATE OF GROWTH? CANCER CELLS DIVIDE in 18 HRS. IS THAT IT?
4 x 18 = 72 hrs. 10 > 20 > 40 > = 80 ?
10) If you wanted to isolate 1 cell out of the suspension culture to start a new suspension culture from a single cell, describe two methods discussed in class to perform this task.
11) What would you predict would happen if you took the initial suspension culture and turned off the mechanism that gently mixed the cells?
12) How would you test your prediction?
13) Assume that you have a cell walking on a substrate. With regard to that cell, describe the various parts of the cells and how these structures give you information about the directionality of the cell. IF I DREW A PICTURE OF THE NON-WALKING CELL TO SHOW NO MOVEMENT - WOULD THIS BE INCORRECT? Or INCORRECT WITHOUT?
14) Describe the process that has to occur in a normal cell if it is to become both anchorage-independent for growth and immotal? I suggest you use a diagram to answer this question.
-----------------------
Outline:
- Intro
- Origin of Cell Biology
- Biomolecular mechanisms / machines
- Problems with Dimenstionality
- Behavior of Cells
- Cell Walking - Culture + in Biology
- Cells as organisms
- How do you look at cells?
------------------------------
Cell Biology
-----------------
It can be seen through 2 historical perspectives
1) Biochemical perspective:
Biochemists were one group of researches who began CB. They homogenized cells first and tested these parts of cells
2) Morphologists:
Used ELETRON MICROSCOPY - they fixed and viewed cells under a microscope to find out more about them.
In the 1950's CB had groups involved like the (ASM) Microbiologists - wanted to understand "how cells work"
--They were trying to understand what cells DO. Cells were NOT intuitive - scientists didn't know how they worked
"Evolution designed cells" - if cells had desirable characteristics, they lived. If they didn't, they died
What are "Biomolecular machines" and what do they do? How do we understand biomolecular machines?
First approach is DESCRIPTIVE:
If we want to describe anything, what do we do? We look at it, look at its structures. Macroscopic observation means looking at something overall and describing what we know empirically by observing with our eyes, senses.
BUT - this doesn't work with CELLS. Why not? Not because:
- Cells are small, TINY - we can't actually see them - so we have to use MACHINES to do this.
- Electron Microscopy is a technology we use - BUT - our view are only FREEZE FRAME picutres
- It took 40+ years to figure out some basic structures.
SO - Cells are THEORETICAL because of out limits of observation
Given all this, since we have to have indirect approaches to study cells - theoretical ways - scientists came up with approaches to do so.
Cliques arose between the BIOCHEMISTS and MICROBIOLOGISTS camps and the didn't agree upon which approach was best - do we study the minute details - the individual pieces because we couldn't look at the parts in an intact cells, or did we look without a lot of detail at whole cells?
- Example: Microfilaments, a basic structure in cells - was huge source of contention. I remember going to a conference where grown men called each other names and used four-letter words over whether microfilaments exisited. MIC's insisted that they could look right at cells and see them. BCHs laughed and said "you're crazy"- when we break the cells up, there is absolutely no evidence they exist.
Biochemists like to isolate single, pure parts of a cells or molecules and study, test these. They take a tube with a receptacle and a plunger, and plunge, plunge to break cells apart and then centifuge. At the bottom of the tube you get "gemisch" and plasma above that. So they take these cells parts from the bottom and study them. What might be a limitation for this? It's one approach, BUT - "all cells come from cells". When you do this - are you studying a live cell any more? Can you tell exactly what it's doing? Can you tell by observing - what the cell is doing? NO - it's DEAD!
REVIEW THIS - CAN YOU CLARIFY WHAT YOU MEANT BY THE LIGHT SWITCH and MARKERS?
So - to see a live, or at least entatct, cell, you put it under a Microscope, you look at it. What do you see [refers to a drawing on board] let's say we have these structures and parts. What is "uniform" vs. "non-uniform" - what are separate distinct stuctures? What do they do? We cam to realize over time that these were separate "machines" with specific functions.
Example: let's say you come into a room you've never been in before. You see the light switch. Even if you've never seen one like this before, could you guess at what is does? Could you guess that it at least does something specific because it is discrete and it's there? You walked into the room, and you turned it on. It is distinct and has a function. You did these in sequence. The same with these board markers.
This is the same thing with cells. We look at cells and we can tell that because there are structures, organization, distinct or discrete characteristics, that these parts and stuctures have function. We may not know what they do - we still don't know what some of these things do - but we know that the cells have evolved to have them as "machines".
This is the same as biotechnology - they take these machines, the mechanisms of cells and do tasks - like the light switch - ON DEMAND and OUT OF SEQUENCE, to meet the needs the researchers are looking for.
DIMENSIONALITY
-----------------------------
WHICH EM IS THE BEST? SEM? TEM?
- Cells are SMALL
- Our understanding of cells is ABSTRACT
- ELECTRON MICROSCOPY is in STILL FRAMES (DEAD CELLS)
- Thin 2-D slices is what we see in Trasnmission Electron Microscopy. So it would be like you looked at me through thin slices - would you be able to get a good idea of how I looked if this is what you saw?
- Cytologic FIXATION KILLS Cell
- Scanning Electon Microscopy is another method that was developed and it addressed some of the drawbacks of TEM - it shows 3-D views of cells at the surface but at one time was not as high resolution as now. SEM is better, higher resolution than LIGHT MICROSCOPY - you can see filaments such as FILIPODIA which are only 0.2 x 10 micrometers - much smaller structures than can be seen with LM. So - SEM has about a 0.2 micrometer resolution.
Drawbacks to SEM:
- Can't see within cells
- Can't see motion
LIGHT MICROSCOPY (LIVING CELLS):
- Bright field - use stained slides
- Phase constast
- Differential Interferometry?!?! Contrast
---------------------------------------------------------------
CELL BEHAVIOR - phenomenological behavior, involves how cells are predicted to act, based on what we've observed of their behavior. For example, if I see a cell moving and it stops, I can predict with good accuracy what the cell will do next.
So - if we drop cells into a culture, and they are suspended and free to move about - what shape will they assume?
- SPHERE!!! Why is this? Because the sphere is a shape has the smallest sruface area with the largest volume. All cells in culture assuma SPHERICAL shape and fall to the bottom of their container. What's interesting is that when they hit that bottom, they being to explore. Let me explain....
First, though - tell me, what are some examples of cell types? [From stus: hepatocytes, hempcytes. These react DIFFERENTLY to their enrivonment. For example - if I put a rabbit into a box - what happens? It will go to the middle of the box. If I do the same with a rat - what do you predict will happen? It will run to the corner and cower. These are different species, but just by knowing what they are and having observed them, I can predict with great certainty what they will do]
So - we have cells, say our heptocytes that hit the botton of the container - what do they do? They are sphereical as they fall - but look at the picture [Has a Sphere, half-sphere, half-flat, and somewhat flat cells]
They touch the bottom, and:
FILIPODIA: - the cell initially extends these in all directions. They are tiny hair-like, like fingers to "feel" their environment. 0.2 micometers in diameter, 2-10 micrometers in length. So, the cell extends these out to feel. As it does this, the cell begins to touch the bottom of the container. The cells does this for a while and then stops 2-3 minutes - a pause. Then...
LAMELLIPODIA: once the cell senses the surface with FILIPODIA and pauses, it extends a larger stucture like a leg, engulfing "swallowing" the FILIPODIA as it extends. As the cell extends a LAMELLIPODIA, is secretes a substances onto the surface of the container - the "cellular equivalent of superglue" so that it sticks and continues extending until FLAT and like a pancake. It may stay there, or it may begin to "WALK". [Note the drawings of a "pancake" with a hump in the middle - NONW WALKING and one with the hump to a side - WALKING.] The lamellipodia then assembles actin and miosin filaments together to form the "cellular equivalent of a muscle" which contracts to MOVE THE CELL?!?!? then sends another lamellipodia out in the direction it wants to go. It will only do this in the direction of the surface of the container. The cell won't waste ATP trying to extend lamellipodia in a diection it won't be able to move in.
As the cell contracts and WALKS, it leaves behind RETRACTION FIBERS - which are the superglue form where the lamellipodia attached and small blebs of plasma membrane that trail behind. Eventually these release and merge (mostly) back into the cell's membrane and cytoplasm. After the retraction fiber releases, there will still be traces of superglue and membrane left - MICROEXUDATES.
As the cell moves, it "drags" the Nucleus (which causes the bulge) using ATP ONLY in the lamellipodia. (The cell doesn't waste ATP by forming and contracting "muscles" in the part of the cell around the trailing Nucleus.)
posted by Ari at 9:55 PM
Saturday, July 09, 2005
Lecture 4
What's cell inhibition?
Cell hits another, it recoils or is reflected and changes direction, possibly away in the angle it came in.
Tell me about Anchorage dependence for growth? What has to happen for a normal cell to be arrested in the cell cycle? It becomes unanchored. 1-2 ft - 50% unchachored. Different levels of anchoredness. Pancake shaped (most highly anchored), a little more round, until touching only in one point.
Judah Folkman - tested, stepwise, using liquid teflon. Made a whole plate of cells that were all the same shape or certain roundness. cells changing from a "pancake" shape to touching only in one point. At a certain % roundedness, protein synthesis stops. mRNA continues, but after more roundedness, mRNA synthesis stops. A little more roundness, cell still attemps to go thru cell cycle, until only a teeny amount of anchorage is left, then even the cell cycle - DNA syntesis - stops. So - know that as cells moves from pancake to unattached (sphere), in stages it stops going thru the cell cycle. AD is the ability for cells to inhibit going thru the cell cycle as they become more round, which isn't the complete loss of anchorage but reduced anchorage.
Cancer cells are AID - will grow even if only attached at one tiny point, or even in suspension. ANS CC's do NOT display contact inhibition.
IMMORTAL MEANS: given enough nutrient and given surface area to grow in - they will grow forever and ever. This is definintive - we know from the experiment from a cell line that has grown continuously since 1952-3. First cell to grow a long long time was taken from (Henrietta Lacks , a black woman). White Dr.s got permission, but didn't tell her all the research done to them. Granddaughter asked to be acknoledged a few years ago, Dr.s resisted at first.
Helped the polio vaccine, etc. - many parts of research.
Immortal is 1/2 way to becoming a cancer cell.
Time line - human fibroblast (from a foreskin) grow in culture for an absolute number of times 16 gens. in culture.
Cells can be
Cell hits another, it recoils or is reflected and changes direction, possibly away in the angle it came in.
Tell me about Anchorage dependence for growth? What has to happen for a normal cell to be arrested in the cell cycle? It becomes unanchored. 1-2 ft - 50% unchachored. Different levels of anchoredness. Pancake shaped (most highly anchored), a little more round, until touching only in one point.
Judah Folkman - tested, stepwise, using liquid teflon. Made a whole plate of cells that were all the same shape or certain roundness. cells changing from a "pancake" shape to touching only in one point. At a certain % roundedness, protein synthesis stops. mRNA continues, but after more roundedness, mRNA synthesis stops. A little more roundness, cell still attemps to go thru cell cycle, until only a teeny amount of anchorage is left, then even the cell cycle - DNA syntesis - stops. So - know that as cells moves from pancake to unattached (sphere), in stages it stops going thru the cell cycle. AD is the ability for cells to inhibit going thru the cell cycle as they become more round, which isn't the complete loss of anchorage but reduced anchorage.
Cancer cells are AID - will grow even if only attached at one tiny point, or even in suspension. ANS CC's do NOT display contact inhibition.
IMMORTAL MEANS: given enough nutrient and given surface area to grow in - they will grow forever and ever. This is definintive - we know from the experiment from a cell line that has grown continuously since 1952-3. First cell to grow a long long time was taken from (Henrietta Lacks , a black woman). White Dr.s got permission, but didn't tell her all the research done to them. Granddaughter asked to be acknoledged a few years ago, Dr.s resisted at first.
Helped the polio vaccine, etc. - many parts of research.
Immortal is 1/2 way to becoming a cancer cell.
Time line - human fibroblast (from a foreskin) grow in culture for an absolute number of times 16 gens. in culture.
Cells can be
posted by Ari at 10:42 AM
Friday, July 08, 2005
Lecture 3
I have to digest tissue at 37 deg. at 1 hr - but want to have more time, so I do it at 18 deg. and draw it our longer - do it for one hour
Place in sterile plate TWO days, with culture medium present.
Must shake it while heating in incubator. Trypsin Works like butter onto an English muffin - gets trypsin into "nooks and crannies" between cells, shaking helps is get in. As a little "gets in there" - it erodes away the most exposed proteins that link cells together. Might not get 100%, but gets a lot of the linkages.
PRIMARY EXPLANT is all up to step 3. Because we're "explanting" tissue.
"cellular equivalent of superglue"
"cellulat equivalent of a muscle"
Place in sterile plate TWO days, with culture medium present.
Must shake it while heating in incubator. Trypsin Works like butter onto an English muffin - gets trypsin into "nooks and crannies" between cells, shaking helps is get in. As a little "gets in there" - it erodes away the most exposed proteins that link cells together. Might not get 100%, but gets a lot of the linkages.
PRIMARY EXPLANT is all up to step 3. Because we're "explanting" tissue.
"cellular equivalent of superglue"
"cellulat equivalent of a muscle"
posted by Ari at 8:22 AM
