Dear Samset,
I will admit that it has been a long time for me in performing
antibiotic assays.
USP Chapter 81 – Antibiotics-Microbial Assays is a good starting point
for obtaining test organisms, inoculum preparation, preparation of
stock solutions, test dilutions of reference standards, etc.. To
calculate the potency of an antibiotic test sample by using either a
cylinder plate assay or turbidimetric assay, you need to generate a
standard curve by using an antibiotic reference standard. Because you
are using zones of inhibition, I believe that you are performing a
cylinder plate assay. For the cylinder-plate assay, each plate
includes only two treatments, the reference treatment (median level
standard, i.e., S3) and one of the other four concentrations of the
standard (S1, S2, S4, and S5) or the sample (U3). The sample should
be diluted to give a nominal concentration that is estimated to be
equivalent to the median reference concentration (S3) of the standard.
Drop six assay cylinders on the inoculated surface of a Petri dish.
For deriving the standard curve, fill alternate cylinders on each of
three plates with the medium test dilution (S3) of the standard and
each of the remaining nine cylinders with one of the other four test
dilutions of the standard. For the test sample, fill alternate
cylinders on each of three plates with the median test dilution of the
standard (S3), and fill the remaining nine cylinders with the
corresponding test dilution (U3) of the sample. After incubation,
remove the cylinders and measure the diameter of zones of inhibition
to the nearest 0.1 mm. Antibiotic potency for a test sample is
calculated by interpolation from a standard curve using a
log-transformed straight-line method with a least-squares fitting
procedure.
If you are not performing antibiotic assays in-house on a routine
basis, it can be somewhat difficult because it requires a high degree
of expertise that can be lost or degraded if not maintained. As far
as using a contract test laboratory, not all laboratories are
qualified to perform this type of assay. You will need to find a
contract testing laboratory that performs antibiotic assays on a
routine basis. It is very easy to separate the good and bad
laboratories that perform antibiotic potency assays. One way to
separate them is to look at the size of the zones of inhibition for
each of the standard test solutions for an antibiotic that is tested
on different test days. If the sizes of the zone of inhibition for
each of the standard test solutions that are used for generating the
standard curve are identical in size to data from other test days, it
shows that short-cuts are being used to perform antibiotic assays.
These zones of inhibition for a standard curve should be similar in
size but not exactly identical in size over a period of time.
Don
Donald J. English Microbiological Quality Consulting LLC
Florham Park, New Jersey 07932
On Tue, Mar 14, 2023 at 11:19 AM Tony Cundell <[log in to unmask]> wrote:
>
> I would refer you to USP <81> Antibiotics - Microbial Assays
>
> I would caution you microbial assays require much skill and experience so I
> recommend you employ a contact testing lab
>
> On Tue, Mar 14, 2023 at 10:10 AM Samet Sarıca <[log in to unmask]> wrote:
>
> > Hello All
> >
> > I need help microbial assay analyzes calculation. How can i calculate
> > results of zones in petri dishes.
> >
> > Regards
> >
> > Samet
> >
> > iPhone’umdan gönderildi
> >
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