I have 100 mg of peptide antibiotic i.e colistin. Potency is >15000 U/mg and solubility is in water (50 mg/ml according to manufacturer's instructions). As potency is given in units, so converting (via online conversion calculators) it into micrograms, it is 1U=731.7 ug in case of colistin. As W=C*V/P formula does not apply on peptide antibiotics. So, how should the stock solution be made if my final testing concentration is 128 mg/L but first I need to make 20 X of this concentration. (128 mg/L would be the concentration in 150 ul volume). I'll appreciate if anyone can help in this.
Hi there,
No need of potency here as you intend to assay mass concentrations (btw there must be something wrong in your calculation as your calculated unit is uncompatible with the potency...). 20x stock should be 2.56mg/mL. Te important thing to know about the starting material is it's actual content in colistin (when pure it's 30000U/mg).
Hi there,
No need of potency here as you intend to assay mass concentrations (btw there must be something wrong in your calculation as your calculated unit is uncompatible with the potency...). 20x stock should be 2.56mg/mL. Te important thing to know about the starting material is it's actual content in colistin (when pure it's 30000U/mg).
My colleague from Thailand worked with colistin last year. He stated how colistin stock solution should be made very quickly, because the powder can vaporize easily.
Thank you so much for your replies.
@Dominique Liger
So, it means I just weigh it directly without going into potency. As you mentioned 2.56 mg/ml would be my 20 X.
Kindly clear another confusion. According to protocol,I'll dilute this 20 X by 1:10 in Muller hinton broth, let's say I make 100 ul of this and add it in first well of my 96 well plate along with 50 ul of media and serially dilute it further up to to 10 dilutions in 96 well plate. In the end, I'll be adding 50 ul of bacteria to each well as well. So it would be 150 ul of volume in each well. My testing concentration remains 128 mg/L in first well? So, I am doing it fine, right?
Hi again,
If final volume is 200µL (100µL of 1/10th dilution of stock plus 50µL of medium plus 50µL of cells) then final dilution is actually 1/20th which is what you were looking for for the first well, ie.128µg/mL (be careful as the dilution factor would only be 1/15th if the final volume is 150µL). For further dilutions, start from the 1/10th dilution and prepare each well accordingly by mixing 100µL of the dilution with 50µL of media and 50µL of cells.
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