Biomarkers and Precision Medicine Community (BPMC)

A recent discussion came up on biomarker parallelism and was wondering what has been the standard practice in the industry. Do you perform parallelism with neat sample even though the kit recommends a DF or has an MRD? 

For example, if the kit recommends a 100 fold dilution, do you perform parallelism at DF=1, 2, so on or at DF=100, 200, ..? In another example, if the STDs, QCs and samples have a MRD of 5, do you perform parallelism at DF=1, 2, so on or DF=5, 10,..?

Thank you in advance for your response!

I recommend you consider performing serial dilution of the test sample using a well-characterized matrix pool (if feasible) and then performing the 1:100 MRD (with buffer) on the diluted samples.  In this way the % matrix will remain constant across all samples.  This approach will only work if the biomarker-of-interest in the original test sample is present at a sufficiently high concentration to generate acceptable interpolated results after dilution.

A recent discussion came up on biomarker parallelism and was wondering what has been the standard practice in the industry. Do you perform parallelism with neat sample even though the kit recommends a DF or has an MRD? 

For example, if the kit recommends a 100 fold dilution, do you perform parallelism at DF=1, 2, so on or at DF=100, 200, ..? In another example, if the STDs, QCs and samples have a MRD of 5, do you perform parallelism at DF=1, 2, so on or DF=5, 10,..?

Thank you in advance for your response!

I fully support and agree with Lyndsay's post in reply. Additionally, the practice is well and most recently described and recommended by consensus in the C-Path paper - Points to Consider Document: Scientific and Regulatory Considerations for the Analytical Validation of Assays Used in the Qualification of Biomarkers in Biological Matrices. (2019)

There is a future paper in preparation being lead by Lyndsay for the AAPS.

What is described step-by-step by C-Path will almost universally be applicable. Don't try and make it more complicated if this produces satisfactory results would be my advice - and remember the actual definition of parallelism:-

In simple terms, Parallelism can be described as recovery in dilution of the endogenous biomarker of interest in the biological matrix of interest. Hence, parallelism is a condition in which dilution of test samples does not result in biased measurements of analyte concentration. Thus, when a test sample is serially diluted to result in a set of samples having analyte concentrations that fall within the quantitative range of the assay, there is no apparent trend towards increasing or decreasing estimates of analyte concentrations over the range of dilutions.

There may be other considerations depending on results seen and some of these may include a look at admixing (what Ron may be suggesting in part I think?)  but the ways to do that are usually considered case by case for specific biomarkers and circumstances. Having worked in both diagnostic and research developments of Biomarker assays for over 50 years now, I've rarely needed to do this and so I won;t complicate the issue here. 

I recommend you consider performing serial dilution of the test sample using a well-characterized matrix pool (if feasible) and then performing the 1:100 MRD (with buffer) on the diluted samples.  In this way the % matrix will remain constant across all samples.  This approach will only work if the biomarker-of-interest in the original test sample is present at a sufficiently high concentration to generate acceptable interpolated results after dilution.

A recent discussion came up on biomarker parallelism and was wondering what has been the standard practice in the industry. Do you perform parallelism with neat sample even though the kit recommends a DF or has an MRD? 

For example, if the kit recommends a 100 fold dilution, do you perform parallelism at DF=1, 2, so on or at DF=100, 200, ..? In another example, if the STDs, QCs and samples have a MRD of 5, do you perform parallelism at DF=1, 2, so on or DF=5, 10,..?

Thank you in advance for your response!

I fully support and agree with Lyndsay's post in reply. Additionally, the practice is well and most recently described and recommended by consensus in the C-Path paper - Points to Consider Document: Scientific and Regulatory Considerations for the Analytical Validation of Assays Used in the Qualification of Biomarkers in Biological Matrices. (2019)

There is a future paper in preparation being lead by Lyndsay for the AAPS.

What is described step-by-step by C-Path will almost universally be applicable. Don't try and make it more complicated if this produces satisfactory results would be my advice - and remember the actual definition of parallelism:-

In simple terms, Parallelism can be described as recovery in dilution of the endogenous biomarker of interest in the biological matrix of interest. Hence, parallelism is a condition in which dilution of test samples does not result in biased measurements of analyte concentration. Thus, when a test sample is serially diluted to result in a set of samples having analyte concentrations that fall within the quantitative range of the assay, there is no apparent trend towards increasing or decreasing estimates of analyte concentrations over the range of dilutions.

There may be other considerations depending on results seen and some of these may include a look at admixing (what Ron may be suggesting in part I think?)  but the ways to do that are usually considered case by case for specific biomarkers and circumstances. Having worked in both diagnostic and research developments of Biomarker assays for over 50 years now, I've rarely needed to do this and so I won;t complicate the issue here. 

I recommend you consider performing serial dilution of the test sample using a well-characterized matrix pool (if feasible) and then performing the 1:100 MRD (with buffer) on the diluted samples.  In this way the % matrix will remain constant across all samples.  This approach will only work if the biomarker-of-interest in the original test sample is present at a sufficiently high concentration to generate acceptable interpolated results after dilution.

A recent discussion came up on biomarker parallelism and was wondering what has been the standard practice in the industry. Do you perform parallelism with neat sample even though the kit recommends a DF or has an MRD? 

For example, if the kit recommends a 100 fold dilution, do you perform parallelism at DF=1, 2, so on or at DF=100, 200, ..? In another example, if the STDs, QCs and samples have a MRD of 5, do you perform parallelism at DF=1, 2, so on or DF=5, 10,..?

Thank you in advance for your response!

A recent discussion came up on biomarker parallelism and was wondering what has been the standard practice in the industry. Do you perform parallelism with neat sample even though the kit recommends a DF or has an MRD? 

For example, if the kit recommends a 100 fold dilution, do you perform parallelism at DF=1, 2, so on or at DF=100, 200, ..? In another example, if the STDs, QCs and samples have a MRD of 5, do you perform parallelism at DF=1, 2, so on or DF=5, 10,..?

Thank you in advance for your response!