HPLC Validation - Calibration Curve Help

[hello345]

Hi Guys

I am currently developing an ion chromatography method for the simultaneous analysis of Cl, Br and I. This is my first time performing a proper validation and I have a question regarding calibration.

From the guidelines I have been following to perform my validation, it states that 'the blank matrix sample should not be taken into consideration when plotting calibration curves'. I was wondering wether this approach was taken by others who have previous experience of method validation? How do you construct your graph i.e. force point through zero? plot blank sample? only plot validation standards minus the blank sample?

Thank you for your help in advance.

[tom jupille]

force point through zero?

No

plot blank sample?

No

only plot validation standards minus the blank sample?

Yes

[hello345]

Hi Tom

Thank you for your reply - it is much appreciated.

Could I please have some clarification on the 'only plot validation standards minus the blank sample' statement I made? I dont think I worded it very well.

Does this mean:

(A) I only plot the calibration standards
(B) I only plot the calibration standards (after subtracting any blank matrix areas from the calibration standards areas)

Thank You

[tom jupille]

(A) I only plot the calibration standards

[BHolmes]

I would strongly suggest that your calibration standards match the matrix of your samples. Use a clean sample to make calibration standards and also inject it as a matrix blank.

I don't know what your application is but from my experience with chromatography, a calibration curve should always be made using matrix matched standards.

[tom jupille]

a calibration curve should always be made using matrix matched standards.

I agree, but that wasn't the question. The question was interpretation of the statement 'the blank matrix sample should not be taken into consideration when plotting calibration curves'

[Jru]

For what it's worth, we also don't have a calibration blank in our curve. Some methods require a force through zero but other than that we don't. Only the standards themselves are included. The calibration black can be read as a qc standard though.