The Q-Prep system: effects on the apparent expression of leucocyte cell surface antigens.
To facilitate the analysis of immunolabelled peripheral blood or bone marrow leucocytes by flow cytometry, a number of reagents are available commercially that lyse erythrocytes and fix leucocytes. This study has investigated the effect on antibody-labelled whole blood of the Q-Prep procedure, in which erythrocytes are lysed with formic acid, and leucocytes are fixed with formaldehyde. Whole blood samples were labelled with the nuclear dye LDS-751 and with antibodies to HLA-DR or belonging to CD2, CD3, CD4, CD7, CD8, CD10, CD13, CD14, CD19, CD20, CD29, CD33, CD45, CD45RA, CD56, and CD62L (TQ-1) that were directly conjugated to either phycoerythrin (PE) and/or fluorescein isothiocyanate (FITC). Leucocytes were analysed by flow cytometry either in unfixed, unlysed whole blood (15) or after preparation using the Q-Prep system. The binding of eight antibodies, CD19-FITC, CD2-PE, CD3-PE, CD4-PE, CD19-PE, CD29-PE, CD45RA-PE, and CD56-PE, to the surface of lymphocytes was reduced, resulting in significant changes (P < 0.05) in the percentages of cells that stained positively and/or their mean molecules of equivalent fluorochrome (MEF). Further analysis revealed that this was due to the formic acid used during the erythrocyte lysis stage.
Macey
M G
MG
Department of Haematology, Royal London Hospital, Whitechapel, United Kingdom.
McCarthy
D A
DA
Davies
C
C
Newland
A C
AC
D000911:Antibodies, Monoclonal; D015703:Antigens, CD / Q000032:analysis* / Q000276:immunology; D000954:Antigens, Surface / Q000032:analysis* / Q000276:immunology; D004912:Erythrocytes; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D019085:Fluorescent Antibody Technique, Direct; D005561:Formates; D006801:Humans; D016130:Immunophenotyping / Q000379:methods*; D007962:Leukocytes / Q000276:immunology*; D010799:Phycoerythrin
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D000954:Antigens, Surface; D005561:Formates; D010799:Phycoerythrin; D016650:Fluorescein-5-isothiocyanate