Flow cytometric measurement of intracellular bilirubin in human peripheral blood mononuclear cells exposed to unconjugated bilirubin

Yoshio Haga, H. David Kay, Margaret A. Tempero, Rowen K Zetterman

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Human peripheral blood mononuclear cells were incubated at 37 °C with bilirubin in bovine albumin solution. Histological analysis of bilirubin-treated cells demonstrated a prominent brown pigment deposited in the cytoplasm. Homogenates of these cells in chloroform-methanol solution showed an identical absorption spectrum with pure bilirubin dissolved in the same solution. When bilirubin-treated cells were excited at 488 nm, a significant autofluorescence was detected by flow cytometry at 585 nm in a dose-dependent manner, which had a significant correlation with the amount of bilirubin chemically extracted from the cells (r = 0.963, n = 34, p < 0.001). Intraassay and interassay variability of the autofluorescence by flow cytometric analysis was small (both <5%). When bilirubin-treated cells were stained with fluorescein-labeled anti-CD14 antibody, the CD14 positive cell population can be fractionated without interference of autofluorescence derived from intracellular bilirubin. These results suggest that flow cytometric analysis of bilirubin-treated cells can quantitate intracellular bilirubin, and that bilirubin does not interfere with analysis of surface antigens.

Original languageEnglish (US)
Pages (from-to)277-283
Number of pages7
JournalClinical Biochemistry
Volume25
Issue number4
DOIs
StatePublished - Aug 1992

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Flow measurement
Bilirubin
Blood Cells
Blood
Cells
Flow cytometry
Surface Antigens
Chloroform
Fluorescein
Pigments
Methanol
Absorption spectra
Anti-Idiotypic Antibodies
Albumins
Flow Cytometry
Cytoplasm
Antibodies

Keywords

  • jaundice
  • liver disease
  • lymphocyte
  • monocyte

ASJC Scopus subject areas

  • Clinical Biochemistry

Cite this

Flow cytometric measurement of intracellular bilirubin in human peripheral blood mononuclear cells exposed to unconjugated bilirubin. / Haga, Yoshio; David Kay, H.; Tempero, Margaret A.; Zetterman, Rowen K.

In: Clinical Biochemistry, Vol. 25, No. 4, 08.1992, p. 277-283.

Research output: Contribution to journalArticle

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AB - Human peripheral blood mononuclear cells were incubated at 37 °C with bilirubin in bovine albumin solution. Histological analysis of bilirubin-treated cells demonstrated a prominent brown pigment deposited in the cytoplasm. Homogenates of these cells in chloroform-methanol solution showed an identical absorption spectrum with pure bilirubin dissolved in the same solution. When bilirubin-treated cells were excited at 488 nm, a significant autofluorescence was detected by flow cytometry at 585 nm in a dose-dependent manner, which had a significant correlation with the amount of bilirubin chemically extracted from the cells (r = 0.963, n = 34, p < 0.001). Intraassay and interassay variability of the autofluorescence by flow cytometric analysis was small (both <5%). When bilirubin-treated cells were stained with fluorescein-labeled anti-CD14 antibody, the CD14 positive cell population can be fractionated without interference of autofluorescence derived from intracellular bilirubin. These results suggest that flow cytometric analysis of bilirubin-treated cells can quantitate intracellular bilirubin, and that bilirubin does not interfere with analysis of surface antigens.

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