What is a kinase assay and how is it performed?
Q. I have never carried out a kinase assay or been taught about its theory could anyone please explain this to me? I am at university and it has never been included in my course but I came across it in a paper and thought I should find out about it. If anyone can even point me in the direction of a good website that would be great. Thanks.
Asked by Kara P - Fri Mar 13 20:25:12 2009 - - 1 Answers - 0 Comments
A. Kinases are a very important class of enzymes found in the cell. They transfer the outermost phosphate (the gamma-phosphate) from a molecule of ATP onto a serine, threonine, or tyrosine amino acid in a target protein (i.e., kinases "phosphorylate" their substrate proteins). In order to measure this enzymatic activity, scientists often use an in vitro kinase assay. There are a bunch of variations of this technique, but perhaps the most common is as follows: Cells that express the kinase you want to study are lysed with detergent to produce crude cell lysate in a tube. An antibody specific to the kinase you want to study is added to the cell lysate. The antibody will bind to your target kinase. The mixture is incubated with large… [cont.]
Answered by Anand S - Fri Mar 13 21:00:48 2009
Q. I have never carried out a kinase assay or been taught about its theory could anyone please explain this to me? I am at university and it has never been included in my course but I came across it in a paper and thought I should find out about it. If anyone can even point me in the direction of a good website that would be great. Thanks.
Asked by Kara P - Fri Mar 13 20:25:12 2009 - - 1 Answers - 0 Comments
A. Kinases are a very important class of enzymes found in the cell. They transfer the outermost phosphate (the gamma-phosphate) from a molecule of ATP onto a serine, threonine, or tyrosine amino acid in a target protein (i.e., kinases "phosphorylate" their substrate proteins). In order to measure this enzymatic activity, scientists often use an in vitro kinase assay. There are a bunch of variations of this technique, but perhaps the most common is as follows: Cells that express the kinase you want to study are lysed with detergent to produce crude cell lysate in a tube. An antibody specific to the kinase you want to study is added to the cell lysate. The antibody will bind to your target kinase. The mixture is incubated with large… [cont.]
Answered by Anand S - Fri Mar 13 21:00:48 2009
what assay to use to determine a serotype?
Q. which serotype of chlamydia one has for example. what assay can be done to determine it?
Asked by Teez - Fri Apr 3 04:56:51 2009 - - 1 Answers - 0 Comments
A. Plaque assay can be used for infectivity of the different serotypes of chlamydia type I and II. There are many types of tests to detect the different strains, but i think the common ones are: 1. Mouse toxicity prevention test 2. Microinmmunofluorescence test Hope that helps, good luck
Answered by playpwnsu - Fri Apr 3 07:40:18 2009
Q. which serotype of chlamydia one has for example. what assay can be done to determine it?
Asked by Teez - Fri Apr 3 04:56:51 2009 - - 1 Answers - 0 Comments
A. Plaque assay can be used for infectivity of the different serotypes of chlamydia type I and II. There are many types of tests to detect the different strains, but i think the common ones are: 1. Mouse toxicity prevention test 2. Microinmmunofluorescence test Hope that helps, good luck
Answered by playpwnsu - Fri Apr 3 07:40:18 2009
What does it means by R square (R2) value for regression linear in standard curve of protein assay?
Q. I did Bradford assay to get a standard curve in order to determine the protein concentration. And i do get R square (R2)= 0.9789 which is closer to 1. But, i don't understand why it was said that the value closer to 1 is a better indicator to show that my standard curve is good to determine the protein concentration.
Asked by deane - Tue Jul 31 10:46:18 2007 - - 2 Answers - 0 Comments
A. The R^2 value (or the Pearson Coefficient of Determination) is an indicator of how well your data fits a line or curve. The closer the R^2 value is to 1 the better the data fits the curve. In other words, the closer the R^2 value is to 1 the more likely your data points are solutions to the equation that defines your curve. So if you have a line with the equation y=mx+b, the closer your R^2 value is to 1 the more likely that your data points x and y are actual solutions to the equation. This indicates how well your data fits the model you are testing. For a more technical description
Answered by spilzwon - Tue Jul 31 11:02:57 2007
Q. I did Bradford assay to get a standard curve in order to determine the protein concentration. And i do get R square (R2)= 0.9789 which is closer to 1. But, i don't understand why it was said that the value closer to 1 is a better indicator to show that my standard curve is good to determine the protein concentration.
Asked by deane - Tue Jul 31 10:46:18 2007 - - 2 Answers - 0 Comments
A. The R^2 value (or the Pearson Coefficient of Determination) is an indicator of how well your data fits a line or curve. The closer the R^2 value is to 1 the better the data fits the curve. In other words, the closer the R^2 value is to 1 the more likely your data points are solutions to the equation that defines your curve. So if you have a line with the equation y=mx+b, the closer your R^2 value is to 1 the more likely that your data points x and y are actual solutions to the equation. This indicates how well your data fits the model you are testing. For a more technical description
Answered by spilzwon - Tue Jul 31 11:02:57 2007
What is a membrane fragility assay? What's the significance of the result for this assay? ?
Q. What is a membrane fragility assay? What's the significance of the result for this assay? thanks
Asked by student - Tue Sep 9 10:42:49 2008 - - 1 Answers - 0 Comments
A. The Cell membrane generally constitutes an initial line of protection for the cell and hence for the entire organism for that matter. This basic tennet underlines the various interests and hitech methods for evaluating the intrinsic membrane characteristics: its stability, fluidity, deformability and other viscoelastic properties . Extended to the human red blood cells (RBC) specific alteration of any of these factors are frequently implicated in many physiological and pathophysiological processes, including various erythrocyte based hereditary disorders Estimation of the osmotic fragility (OF) of RBCs for example exploits the specific structural changes which the RBC membrane undergoes when the cell is subjected to osmotic stress Properly… [cont.]
Answered by Peter S - Sat Sep 13 06:46:39 2008
Q. What is a membrane fragility assay? What's the significance of the result for this assay? thanks
Asked by student - Tue Sep 9 10:42:49 2008 - - 1 Answers - 0 Comments
A. The Cell membrane generally constitutes an initial line of protection for the cell and hence for the entire organism for that matter. This basic tennet underlines the various interests and hitech methods for evaluating the intrinsic membrane characteristics: its stability, fluidity, deformability and other viscoelastic properties . Extended to the human red blood cells (RBC) specific alteration of any of these factors are frequently implicated in many physiological and pathophysiological processes, including various erythrocyte based hereditary disorders Estimation of the osmotic fragility (OF) of RBCs for example exploits the specific structural changes which the RBC membrane undergoes when the cell is subjected to osmotic stress Properly… [cont.]
Answered by Peter S - Sat Sep 13 06:46:39 2008
Bradford Dye Method of protein assay be considered a destructive, or non destructive method?
Q. Would the Bradford Dye Method of protein assay be considered a destructive, or non destructive method of analysis? Explain.
Asked by blazey - Tue Mar 17 21:46:19 2009 - - 1 Answers - 0 Comments
A. Can you recover the protein intact after doing the assay? That should answer your question.
Answered by Yuu - Thu Mar 19 18:06:09 2009
Q. Would the Bradford Dye Method of protein assay be considered a destructive, or non destructive method of analysis? Explain.
Asked by blazey - Tue Mar 17 21:46:19 2009 - - 1 Answers - 0 Comments
A. Can you recover the protein intact after doing the assay? That should answer your question.
Answered by Yuu - Thu Mar 19 18:06:09 2009
What is a destructive and nondestructive assay?
Q. what are some advantages and disadvantages of a destructive and non destructive assay. and what are some examples in each assay?
Asked by amar s - Thu Feb 18 00:43:18 2010 - - 0 Answers - 0 Comments
Q. what are some advantages and disadvantages of a destructive and non destructive assay. and what are some examples in each assay?
Asked by amar s - Thu Feb 18 00:43:18 2010 - - 0 Answers - 0 Comments
where can i find a method to assay an antibiotic?
Q. can anyone tell me a method to assay an antibiotic and/or an antiseptic? or send me a link? thanks
Asked by Elle - Mon Feb 8 13:47:36 2010 - - 1 Answers - 0 Comments
A. Which one? What concentration range in what kind of sample? The procedure will be different if you're testing that a tablet is 99.% pure antibiotic, versus measuring 100 ppb levels of antiseptic in waste water. The generic approach is some kind of chromatography - either HPLC or GC, depending on the sample, concentration, the kind of detector you want to use, and whether the antibiotic is stable at high temperature (many aren't). You'd need a small amount of the pure material to prepare standards (so that you know which peak in the chromatogram corresponds to the antibiotic you're looking for, and so that you know how much peak area corresponds to how much mass or concentration of antibiotic). Depending on which antibiotic you're… [cont.]
Answered by Mike S - Mon Feb 8 16:01:30 2010
Q. can anyone tell me a method to assay an antibiotic and/or an antiseptic? or send me a link? thanks
Asked by Elle - Mon Feb 8 13:47:36 2010 - - 1 Answers - 0 Comments
A. Which one? What concentration range in what kind of sample? The procedure will be different if you're testing that a tablet is 99.% pure antibiotic, versus measuring 100 ppb levels of antiseptic in waste water. The generic approach is some kind of chromatography - either HPLC or GC, depending on the sample, concentration, the kind of detector you want to use, and whether the antibiotic is stable at high temperature (many aren't). You'd need a small amount of the pure material to prepare standards (so that you know which peak in the chromatogram corresponds to the antibiotic you're looking for, and so that you know how much peak area corresponds to how much mass or concentration of antibiotic). Depending on which antibiotic you're… [cont.]
Answered by Mike S - Mon Feb 8 16:01:30 2010
what is the principle of the DPPH free radical inhibition assay?
Q. and what has the DPPH assay got to do with absorbance and percent inhibition? i dont know how to interpret my results. and oh yeah i need sources :)
Asked by marketer - Sun Mar 1 12:10:59 2009 - - 1 Answers - 0 Comments
A. Read that website
Answered by adel a - Wed Mar 4 20:14:04 2009
Q. and what has the DPPH assay got to do with absorbance and percent inhibition? i dont know how to interpret my results. and oh yeah i need sources :)
Asked by marketer - Sun Mar 1 12:10:59 2009 - - 1 Answers - 0 Comments
A. Read that website
Answered by adel a - Wed Mar 4 20:14:04 2009
How does methylation interference assay works?
Q. I'm reading a paper where they used this assay to determine which nucleotides participate in binding of a transcription factor. I, however, have not previously learned about this assay and don't seem to find a good description and explanation of its principles online. Can you explain in or direct me to a site where it is explained?
Asked by Kaytee - Mon Feb 26 14:46:21 2007 - - 1 Answers - 0 Comments
A.
Answered by ger - Mon Mar 5 16:01:58 2007
Q. I'm reading a paper where they used this assay to determine which nucleotides participate in binding of a transcription factor. I, however, have not previously learned about this assay and don't seem to find a good description and explanation of its principles online. Can you explain in or direct me to a site where it is explained?
Asked by Kaytee - Mon Feb 26 14:46:21 2007 - - 1 Answers - 0 Comments
A.
Answered by ger - Mon Mar 5 16:01:58 2007
Is there a simple assay that could test the antioxidant content level in various foods?
Q. I want to perform a science fair project on the possible differing antioxidant content levels in fruits and vegetables before and after being cooked by various methods, but do not know how to test for the antioxidant levels in a relatively cheap and simple way.
Asked by Lynn - Sun Dec 20 02:52:08 2009 - - 1 Answers - 0 Comments
A. I'm not sure how cheap it is, but there are simple assays available. I've included links to a couple of relevant articles. You'll want to visit a local university to read these, and may need a science teacher to help translate them. Basically ignore any method that uses GC, MS, HPLC, or any other method that mentions a machine you've never used because chances are you don't have access to such a machine yet. That leaves assays that use a dye or colored substrate, which is bleached by peroxide (oxygen free radicals). The idea is, the more antioxidant there is available, the less oxygen free radical there is to bleach the dye. You will still probably need a spectrophotometer, but every high school biology lab has at least one. DPPH or beta-c [cont.]
Answered by im - Sun Dec 20 04:20:10 2009
Q. I want to perform a science fair project on the possible differing antioxidant content levels in fruits and vegetables before and after being cooked by various methods, but do not know how to test for the antioxidant levels in a relatively cheap and simple way.
Asked by Lynn - Sun Dec 20 02:52:08 2009 - - 1 Answers - 0 Comments
A. I'm not sure how cheap it is, but there are simple assays available. I've included links to a couple of relevant articles. You'll want to visit a local university to read these, and may need a science teacher to help translate them. Basically ignore any method that uses GC, MS, HPLC, or any other method that mentions a machine you've never used because chances are you don't have access to such a machine yet. That leaves assays that use a dye or colored substrate, which is bleached by peroxide (oxygen free radicals). The idea is, the more antioxidant there is available, the less oxygen free radical there is to bleach the dye. You will still probably need a spectrophotometer, but every high school biology lab has at least one. DPPH or beta-c [cont.]
Answered by im - Sun Dec 20 04:20:10 2009
How do you determine protein concentration using the Bradford Assay?
Q. I am really confused, I am supposed to calculate protein concentration from several different tubes containing a protein solution. The units are in ueg/ueL. All I have is the amount of protein concentration for each tube expressed in ueL, and the absorbance of each. How do I find the concentration? I have to make a graph of concentration vs. absorbance.
Asked by Liz - Mon Oct 20 13:43:52 2008 - - 1 Answers - 0 Comments
A. Use this formula to compute for the concentration: concentration = Absorbance (at wavelength used) divided by absorbance coefficient
Answered by sweeter_ion - Tue Oct 21 05:17:31 2008
Q. I am really confused, I am supposed to calculate protein concentration from several different tubes containing a protein solution. The units are in ueg/ueL. All I have is the amount of protein concentration for each tube expressed in ueL, and the absorbance of each. How do I find the concentration? I have to make a graph of concentration vs. absorbance.
Asked by Liz - Mon Oct 20 13:43:52 2008 - - 1 Answers - 0 Comments
A. Use this formula to compute for the concentration: concentration = Absorbance (at wavelength used) divided by absorbance coefficient
Answered by sweeter_ion - Tue Oct 21 05:17:31 2008
Why do we need to dilute proteins for the Bradford Assay?
Q. What is the reason we dilute the proteins and not just use the original cell fractions? Do we need a solution that has more or less protein in it to get a good absorbance?
Asked by ruz - Mon Jul 6 15:25:29 2009 - - 3 Answers - 0 Comments
A. Beer's Law breaks down at high concentrations. Dilute samples give a reaction on the linear portion of the curve. One of the disadvantages of the assay is that it has only a fifty- to one-hundred-fold working range.
Answered by novangelis - Mon Jul 6 15:35:37 2009
Q. What is the reason we dilute the proteins and not just use the original cell fractions? Do we need a solution that has more or less protein in it to get a good absorbance?
Asked by ruz - Mon Jul 6 15:25:29 2009 - - 3 Answers - 0 Comments
A. Beer's Law breaks down at high concentrations. Dilute samples give a reaction on the linear portion of the curve. One of the disadvantages of the assay is that it has only a fifty- to one-hundred-fold working range.
Answered by novangelis - Mon Jul 6 15:35:37 2009
You are going to do an enzyme assay, and you need to make a solution of substrate.?
Q. The concentration of substrate that you will use is 0.001 ug/ml. It is necessary to have enough for the enzyme assay. You will need to test 50 samples, plus a negative control lacking the enzyme, and another negative control lacking the substrate. You have determined that you will do all of this in triplicate. The assay calls for 1 ml of substrate solution and 0.2 ml of enzyme solution. If you don't have enough substrate solution, you will have to make more in the middle of the experiment, which could ruin your results. a. How much substrate solution should you make? b. How many grams of substrate will you need for this solution? c. How can you make this solution if you don't have a balance accurate enough to weigh such a small quantity? I… [cont.]
Asked by Lena - Fri Feb 23 07:16:10 2007 - - 1 Answers - 0 Comments
A. a. How much substrate solution should you make? 50 samples + a negative control (do all of this in triplicate) The assay calls for 1 ml of substrate solution Therefore, (50 + 1)x1 ml x 3 = 153 ml b. How many grams of substrate will you need for this solution? concentration of substrate that you will use is 0.001 ug/ml Therefore, 153 ml x 0.001 ug/ml = 0.153 ug c. How can you make this solution if you don't have a balance accurate enough to weigh such a small quantity? Weigh 1.0mg of substrate and dissolve it in 1000 ml buffer solution in a volumetric flask (stock solution). The resulting concentration will be 1.0 mg/1000ml = 0.001mg/mL (i.e. 1 ug/ml ) Take 1ml from stock solution and dilute it to 1000 ml with buffer solution in a… [cont.]
Answered by Yy - Fri Feb 23 10:03:34 2007
Q. The concentration of substrate that you will use is 0.001 ug/ml. It is necessary to have enough for the enzyme assay. You will need to test 50 samples, plus a negative control lacking the enzyme, and another negative control lacking the substrate. You have determined that you will do all of this in triplicate. The assay calls for 1 ml of substrate solution and 0.2 ml of enzyme solution. If you don't have enough substrate solution, you will have to make more in the middle of the experiment, which could ruin your results. a. How much substrate solution should you make? b. How many grams of substrate will you need for this solution? c. How can you make this solution if you don't have a balance accurate enough to weigh such a small quantity? I… [cont.]
Asked by Lena - Fri Feb 23 07:16:10 2007 - - 1 Answers - 0 Comments
A. a. How much substrate solution should you make? 50 samples + a negative control (do all of this in triplicate) The assay calls for 1 ml of substrate solution Therefore, (50 + 1)x1 ml x 3 = 153 ml b. How many grams of substrate will you need for this solution? concentration of substrate that you will use is 0.001 ug/ml Therefore, 153 ml x 0.001 ug/ml = 0.153 ug c. How can you make this solution if you don't have a balance accurate enough to weigh such a small quantity? Weigh 1.0mg of substrate and dissolve it in 1000 ml buffer solution in a volumetric flask (stock solution). The resulting concentration will be 1.0 mg/1000ml = 0.001mg/mL (i.e. 1 ug/ml ) Take 1ml from stock solution and dilute it to 1000 ml with buffer solution in a… [cont.]
Answered by Yy - Fri Feb 23 10:03:34 2007
Calculating concentration of NADH and pyruvate before the start of an assay?
Q. 0.1ml NADH and 0.01ml pyruvate in the assay. were given 3.4mM NADH and 30mM so what would the concentrations be in the assay?
Asked by Gemma N - Tue May 29 17:40:40 2007 - - 1 Answers - 0 Comments
A. Data required.
Answered by ag_iitkgp - Wed May 30 12:25:18 2007
Q. 0.1ml NADH and 0.01ml pyruvate in the assay. were given 3.4mM NADH and 30mM so what would the concentrations be in the assay?
Asked by Gemma N - Tue May 29 17:40:40 2007 - - 1 Answers - 0 Comments
A. Data required.
Answered by ag_iitkgp - Wed May 30 12:25:18 2007
How do you know which type of viral assay method to use for a particular virus? (Hemagluttination assay etc.)?
Q. What are the purposes of different assays?
Asked by Matt - Fri Oct 23 16:19:51 2009 - - 1 Answers - 0 Comments
A. Different viruses have different characteristics that require different methods. Also, what you are testing will determine what kind of assay you need. If there was only one assay, only one virus could be tested for one thing.
Answered by unknown - Fri Oct 23 16:38:38 2009
Q. What are the purposes of different assays?
Asked by Matt - Fri Oct 23 16:19:51 2009 - - 1 Answers - 0 Comments
A. Different viruses have different characteristics that require different methods. Also, what you are testing will determine what kind of assay you need. If there was only one assay, only one virus could be tested for one thing.
Answered by unknown - Fri Oct 23 16:38:38 2009
How do you calculate for the concentration of two unknown absorbances in Bradford assay?
Q. Unknown A= 0.359 and Unknown B=0.336. Please help.
Asked by Kathleen H - Sat Jan 2 13:53:36 2010 - - 1 Answers - 0 Comments
Q. Unknown A= 0.359 and Unknown B=0.336. Please help.
Asked by Kathleen H - Sat Jan 2 13:53:36 2010 - - 1 Answers - 0 Comments
How do I propose this cell-free assay?
Q. I hypothesize that cytosolic factors regulate autophagosome formation. How do I set up a cell-free assay to reconstitute autophagosome formation in order to identify these cytosolic regulators? What cell-free assay do I propose to recapitulate cytosolic regulatory activity? (this assay should be autophagy-specific)
Asked by Student - Fri May 15 15:50:56 2009 - - 1 Answers - 0 Comments
Q. I hypothesize that cytosolic factors regulate autophagosome formation. How do I set up a cell-free assay to reconstitute autophagosome formation in order to identify these cytosolic regulators? What cell-free assay do I propose to recapitulate cytosolic regulatory activity? (this assay should be autophagy-specific)
Asked by Student - Fri May 15 15:50:56 2009 - - 1 Answers - 0 Comments
Should you always assume a linear relationship between conc. of DNA and fluorescence in the picogreen assay?
Q. My data for the detection of a DNA plasmid always fits a perfect curve, a linear fit can be assigned to the lower portion of the data. I dont know how to best deal with the data, should I just use the points that may fit linearity or treat the data as a curve? Thanks wise duck, however my data does not plateau as in the case of saturation, I have seen this often in immunoassay, rather it rises exponentially.
Asked by babybaleno - Thu Jan 31 07:52:04 2008 - - 1 Answers - 0 Comments
A. The concentration vs absorbance is only accurate during the linear part of the curve. As you approach saturation the number is unreliable.
Answered by Weise Ente - Thu Jan 31 09:05:29 2008
Q. My data for the detection of a DNA plasmid always fits a perfect curve, a linear fit can be assigned to the lower portion of the data. I dont know how to best deal with the data, should I just use the points that may fit linearity or treat the data as a curve? Thanks wise duck, however my data does not plateau as in the case of saturation, I have seen this often in immunoassay, rather it rises exponentially.
Asked by babybaleno - Thu Jan 31 07:52:04 2008 - - 1 Answers - 0 Comments
A. The concentration vs absorbance is only accurate during the linear part of the curve. As you approach saturation the number is unreliable.
Answered by Weise Ente - Thu Jan 31 09:05:29 2008
What is the purpose of an alkaline buffer in an alakline phosphatase assay?
Q. What is the purpose of an alkaline buffer in an alakline phosphatase assay?
Asked by master - Fri Nov 28 15:01:30 2008 - - 2 Answers - 0 Comments
A. alakline phosphatase will work optimumly at a alkaline pH. pH is a negative log measurment of H+ ions the greater the pH fewer number of H+ ions, if enzymes are out of there optimum range they will either donate or uptake H+ this change in the structure will cahnge the enzymes active site and the enzyme will not be able to catalyze reactions
Answered by Lora - Sun Nov 30 11:10:35 2008
Q. What is the purpose of an alkaline buffer in an alakline phosphatase assay?
Asked by master - Fri Nov 28 15:01:30 2008 - - 2 Answers - 0 Comments
A. alakline phosphatase will work optimumly at a alkaline pH. pH is a negative log measurment of H+ ions the greater the pH fewer number of H+ ions, if enzymes are out of there optimum range they will either donate or uptake H+ this change in the structure will cahnge the enzymes active site and the enzyme will not be able to catalyze reactions
Answered by Lora - Sun Nov 30 11:10:35 2008
Any ideas on designing an assay to measure the amount of protein in samples of cell lysate using small volumes?
Q. Any ideas on designing an assay to measure the amount of protein in samples of cell lysate using small volumes?
Asked by Issy - Wed Feb 11 09:07:52 2009 - - 1 Answers - 0 Comments
A. Spectrophotometry - 280 nm
Answered by donnybrago - Thu Feb 12 11:03:51 2009
Q. Any ideas on designing an assay to measure the amount of protein in samples of cell lysate using small volumes?
Asked by Issy - Wed Feb 11 09:07:52 2009 - - 1 Answers - 0 Comments
A. Spectrophotometry - 280 nm
Answered by donnybrago - Thu Feb 12 11:03:51 2009
From Yahoo Answer Search: 'Assay'
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This pioneering scientific research represents the first example of protein and DNA biomarkers being combined into one assay to optimize performance. ...
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Harriet
Sun, 24 Jan 2010 21:02:00 GM
A Taqman microRNA . assay. was set up using the cDNA created yesterday. I started with 4 different types of cDNA : 1 - HeLa + random hexamers 2 - HeLa + miR-34 primers 3 - SW1353 + random hexamers 4 - SW1353 + miR-34 primers ...
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