Flow Cytometer Performance Characterization, Standardization and Calibration against CD4 on T Lymphocytes Enables Quantification of Biomarker Expressions for Immunological Applications

Published

August 7, 2014

Author(s)

Heba Degheidy, Steven R. Bauer, Gerald Marti, Lili Wang

Abstract

There is an urgent need for developing a procedure for biomarker standardization and quantification in clinical laboratories. Measuring the expression levels of cell antigens is critical for the diagnosis of many diseases, e.g. leukemia, lymphoma and immunodeficiency diseases. One of the most significant challenges in flow cytometry is obtaining inter-laboratory and intra-laboratory consistent and reproducible results across multiple cytometer platforms and locations longitudinally over time. To obtain measurement consistency, the target flow cytometer voltages should be optimized to segregate the negative population from the electronic noise, and to keep the brightest positive population within the dynamic range of each detectors. Then target values should be determined and transferred to selected cytometers. In this study, we optimized a procedure for instrument standardization across three different flow cytometer platforms from the same vendor and in two different locations. The biomarker quantification was implemented on standardized instruments using CD4 expression on T lymphocytes with a known amount of antibody bound per cell as a quantification standard. Our results on blood cell subset typing and CD19 quantification demonstrated that consistent and reliable results could be accomplished between instruments using the developed procedure. Quantitating the expression levels of certain cell biomarkers before, during, and after therapy would provide important information for monitoring antibody-based therapy and could be potentially used to adjust dosing. Presently, we are implementing this protocol to quantify critical disease biomarkers, and making necessary modifications to the procedure to include instruments from different instrument manufacturers.

Citation

Journal of Biomedical Science

Pub Type

Journals

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Keywords

Flow Cytometry, Biomarker quantification, Instrument standardization, Target mean fluorescence intensity (MFI).

Citation

Degheidy, H. , Bauer, S. , Marti, G. and Wang, L. (2014), Flow Cytometer Performance Characterization, Standardization and Calibration against CD4 on T Lymphocytes Enables Quantification of Biomarker Expressions for Immunological Applications, Journal of Biomedical Science, [online], https://tsapps.nist.gov/publication/get_pdf.cfm?pub_id=916136 (Accessed April 19, 2024)