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- It yields 5-10 ug of DNA from a dried specimen swab, instead of 200-500 ng.
- It improves detection sensitivity.
- It's nontoxic.
- Its lysate may be used for PCR without further DNA purification.
- Dried specimen swabs can be shipped and stored long-term at room temperature.
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jessilov20
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In Topic: Quantify gDNA in buccal swab
29 November 2012 - 02:29 PM
DNA binds the cotton swab amazingly tight. Water can only elute ~5% of DNA from a dried specimen swab. We have developed an AquaGenomic methodthat can fully recover the DNA from dried specimen swabs. You can extract the DNA from the buccal swab with AquaGenomic and then quantify the DNA by conventional methods, from UV spec to real-time PCR.
In Topic: low A260/230 problem
27 August 2012 - 04:58 PM
The scans look normal to me. What were the DNA concentrations? Try use 10x higher concentrations.
In Topic: IC50 calculation software (need help)
27 August 2012 - 04:05 PM
Haha... DRT may be right. Or it should read “should never use Excel for IC50 estimation if you can't write a macro that incorporate the four parameter model for nonlinear fitting of a sigmoid dose-response curve.”
DRT, we would be happy to try your Excel IC50 macro if you would post it up, and compare it to BioDataFit, in case BioDataFit goes offline.
DRT, we would be happy to try your Excel IC50 macro if you would post it up, and compare it to BioDataFit, in case BioDataFit goes offline.
In Topic: IC50 calculation software (need help)
25 August 2012 - 10:16 AM
I hope this reply will benefit a lot of students trying to learn how to perform a cytotoxicity experiment and calculate the IC50.
You should never use Excel for IC50 estimation. Your viability data is not linear or fits simple log transformation. For accurate calculation of IC50, a nonlinear curve fitting program that uses the Four Parameter Model is needed.
A paid program such as GraphPad Prims is great, but there is a free online program BioDataFit, which produces the same accuracy as GraphPad and it is easier to use.
To learn how to prepare the raw data for inputting into an IC50 program, read here. To learn what to do if your experimental data does not fit the Four Parameter Model, which can be very frustrating to new users, read here.
Good luck and hope you all will smoothly get your cytotoxicity experiments done and get publication quality IC50s from these tips.
You should never use Excel for IC50 estimation. Your viability data is not linear or fits simple log transformation. For accurate calculation of IC50, a nonlinear curve fitting program that uses the Four Parameter Model is needed.
A paid program such as GraphPad Prims is great, but there is a free online program BioDataFit, which produces the same accuracy as GraphPad and it is easier to use.
To learn how to prepare the raw data for inputting into an IC50 program, read here. To learn what to do if your experimental data does not fit the Four Parameter Model, which can be very frustrating to new users, read here.
Good luck and hope you all will smoothly get your cytotoxicity experiments done and get publication quality IC50s from these tips.
In Topic: Alternative Cytotoxicity Assays
24 August 2012 - 03:14 PM
Obviously viability assays that rely on redox activity are not good in your case. In addition to ATP content assay suggested by Kevin, you may look into Life Tech's CyQUANT® Direct Cell Proliferation Assay that quantitates total DNA in the well or Sigma's
Sulforhodamine B Assay that quantitates intact cells. But both are staining based and I am not sure if your nanoparticles would trap the dyes or not, or if the particles would trap the cell-free DNA or not. If everything fails, maybe you could try EdU or BrdU DNA incorporation based assays.
Sulforhodamine B Assay that quantitates intact cells. But both are staining based and I am not sure if your nanoparticles would trap the dyes or not, or if the particles would trap the cell-free DNA or not. If everything fails, maybe you could try EdU or BrdU DNA incorporation based assays.
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