Document Detail

Near-infrared dyes for six-color immunophenotyping by laser scanning cytometry.
MedLine Citation:
PMID:  12116357     Owner:  NLM     Status:  MEDLINE    
BACKGROUND: To adequately analyze the complexity of the immune system and reduce the required sample volume for immunophenotyping in general, more measurable colors for the discrimination of leukocyte subsets are necessary. Immunophenotyping by the laser scanning cytometer (LSC), a slide-based cytometric technology, combines cell detection based on multiple colors with their subsequent visualization without the need for physical cell sorting. In the present study, the filter setting of the LSC was adapted for the measurement of the far-red emitting dye cyanine 7 (Cy7), thereby increasing the number of measurable commercially available fluorochromes. METHODS: The optical filters of the LSC were replaced-photomultiplier (PMT) 3/allophycocyanin (APC): 740-nm dichroic long pass, and 670-/55-nm bandpass; PMT 4/Cy7: 810-/90-nm bandpass. Peripheral blood leukocytes were stained directly by fluorochrome-labeled antibodies or by indirect staining. The tandem dyes of Cy7 (phycoerythrin [PE]-Cy7, APC-Cy7) and the fluorochromes fluorescein isothiocyanate (FITC), PE, PE-Cy5, and APC were tested alone and in different combinations. RESULTS: With the new filter combination and tandem fluorochromes, Cy7 was measurable at 488-nm (argon laser) or 633-nm (helium-neon laser) excitation. Resolution was in the range of FITC for PE-Cy7 but approximately 30% lower for APC-Cy7; spillover into the respective donor fluorochrome channel for both tandem dyes was prominent. A six-color panel for leukocyte subtyping was designed. CONCLUSIONS: With this adaptation, it is possible to measure the tandem conjugates PE-Cy7 and APC-Cy7. This new setup opens the way for six-color immunophenotyping by LSC.
Andreas O H Gerstner; Dominik Lenz; Wiebke Laffers; Robert A Hoffman; Michael Steinbrecher; Friedrich Bootz; Attila Tárnok
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Publication Detail:
Type:  Journal Article    
Journal Detail:
Title:  Cytometry     Volume:  48     ISSN:  0196-4763     ISO Abbreviation:  Cytometry     Publication Date:  2002 Jul 
Date Detail:
Created Date:  2002-07-12     Completed Date:  2002-12-12     Revised Date:  2006-11-15    
Medline Journal Info:
Nlm Unique ID:  8102328     Medline TA:  Cytometry     Country:  United States    
Other Details:
Languages:  eng     Pagination:  115-23     Citation Subset:  IM    
Copyright Information:
Copyright 2002 Wiley-Liss, Inc.
Department of Otorhinolaryngology/Plastic Surgery, University of Leipzig, Leipzig, Germany.
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MeSH Terms
Fluorescent Dyes*
Image Cytometry / economics,  instrumentation,  methods*
Immunophenotyping / methods*
Lymphocyte Subsets / cytology*
Reproducibility of Results
Reg. No./Substance:
0/Benzothiazoles; 0/Carbocyanines; 0/Fluorescent Dyes; 57282-58-3/cyanine dye 7

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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