Single-color multitarget flow cytometry using monoclonal antibodies labeled with different intensities of the same fluorochrome.

Annals of Laboratory Medicine
Joonhong Park, Kyungja Han

Abstract

We developed a single-color multitarget flow cytometry (SM-FC) assay, a single-tube assay with graded mean fluorescence intensities (MFIs). We evaluated the repeatability of SM-FC, and its correlation with multicolor flow cytometry (MFC), to assess its application as a routine FC assay. We selected CD19, CD3, CD4, and CD8 as antigen targets to analyze a lymphocyte subset. MFIs were graded by adjusting monoclonal antibody (mAb) volumes to detect several cell populations. Dimly labeled mAb was prepared by decreasing mAb volume and the optimum diluted volume was determined by serial dilution. SM-FC repeatability was analyzed 10 times in 2 normal controls. The correlation between SM-FC and MFC was evaluated in 20 normal and 23 patient samples. CV values (0.8-5.0% and 1.3-4.1% in samples 1 and 2, respectively) acquired by SM-FC with CD3-fluorescein α-isothyocyanate (FITC)(dim)+CD4-FITC(bright) and with CD19-FITC(dim)+CD3-FITC(bright) showed good repeatability, comparable to that acquired by MFC (1.6-3.7% and 1.0-4.8% in samples 1 and 2, respectively). Excellent correlation was observed between the 2 methods in the 20 normal samples (B cells, T cells, non-T(helper) cells, and T(helper) cells; r(2)=0.87, 0.97, 0.97, and 0.98, respecti...Continue Reading

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Citations

Jan 1, 2012·Biomicrofluidics·Mariangela MortatoGiuseppe Gigli
Jan 31, 2014·Biological Research for Nursing·Richard R ReichThomas W Klein

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Methods Mentioned

BETA
flow cytometry
blood

Software Mentioned

MedCalc
FACSDiva

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