We are testing virus titer, and have to dilute the virus stock at 1:10,000 (Dilution A) and 1:20,000 (Dilution B) to run qPCR as templates. Then convert the diluted titer to dilution corrected titer. However, the conversion from A or B is not the same value, showing a big difference. I guess the problem is about dilution. Is there any best strategy procedure to complete the dilution within 3-step? Thank you.
One problem with making dilutions is that people often try to use very small volumes like 1 ul to make their dilutions. Pipettors are not that accurate at small volumes. So make a 100-fold dilution (10ul into 1ml) and then again 10ul of that into 1ml and you have a 10,000 fold dilution.
either is fine, the important thing is to be sure the volumes you measure are accurate
Dilution Method 1: 100x100
Suppose stock 10^3 vg/uL
Step 1: Add 10uL into 990uL to make 1:100 dilution
If the error is 1uL, after dilution vg/uL = 11x10^3/1000 = 11.
Step 2: Add 10uL from step 1 into 990uL to make another 1:100.
If error is 1uL, after dilution vg/uL = 11x11/1000 = 0.121
Dilution Method 2: 10x10x10x10
Suppose stock 10^3 vg/uL
Step 1: Add 10uL into 90uL to make 1st 1:10 dilution
If error is 1uL, after dilution vg/uL = 11x10^3/100 = 110
Step 2: Add 10uL into 90uL to make 2nd 1:10 dilution
If error is 1uL, after dilution vg/uL = 11x110/100 = 121
Step 3: Add 10uL into 90uL to make 3rd 1:10 dilution
If error is 1uL, after dilution vg/uL = 11x121/100 = 13.31
Step 4: Add 10uL into 90uL to make 4th 1:10 dilution
If error is 1uL, after dilution vg/uL = 11x13.31/100 = 1.4641
Dilution Method 3: 1x10000
If the error is 1uL,
Add 1uL stock into 10000uL to make 1:10000,
final vg/uL = 2x10^3/10000 =0.2
Dilution Method 3: 1x10000
Suppose no error
Add 1uL stock into 10000uL, final vg/uL = 1x10^3/10000 = 0.1
Conclusion: 100x100 is the best if the pipetting error is 1uL for a 10uL tip.
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