Allo-7: a new fluorescent tandem dye for use in flow cytometry

Cytometry. 1996 Aug 1;24(4):390-5. doi: 10.1002/(SICI)1097-0320(19960801)24:4<390::AID-CYTO11>3.0.CO;2-K.


This report describes the development of a novel tandem dye by combining allophycocyanine (APC) and cyanine dye indotricarbocyanine (CY7) to create ALLO-7 for use in flow cytometry. The APC donor fluorophore was excited at 647 nm and, through resonance energy transfer to the CY7 acceptor, produced fluorescence at > 780 nm. To test the applicability of this tandem in single and multicolor immunofluorescence, a streptavidin conjugate of the tandem (SA-ALLO-7) was used for the detection of cell surface antigens on human peripheral blood leukocytes (PBL) by indirect immunofluorescence. Human PBL were stained with CD4/ GaM-APC, CD3-fluorescein isothiocyanate (FITC), CD14-phycoerythrin (PE), CD19-energy-coupled dye (phycoerythrin-Texas Red) (ECD), and CD8-biotin with SA-ALLO-7 and analyzed for fluorescence on a FACS Vantage using dual-laser excitation (488 and 647 nm). The results indicated that the percentage of cells positive for each of the surface antigens was comparable for single-color controls and multicolor samples. The ALLO-7 fluorescence, which was collected with a 730-shortpass dichroic mirror and a 790/50-bandpass filter, was clearly resolved from the APC fluorescence and that from FITC, PE, and ECD. The SA-ALLO-7 exhibited minimal nonspecific binding to PBL monocytes. However, the specific binding of the tandem to high-density antigens was clearly identified by positive fluorescence. This unique tandem reagent, ALLO-7, provided the capability for dual-color immunofluorescence with a 647-nm laser line (or a helium neon laser at 633 nm) and provides the potential to perform three-color analysis with a dye-head laser (Texas Red, APC, ALLO-7).

MeSH terms

  • Antibodies, Monoclonal
  • Carbocyanines / chemistry*
  • Coloring Agents / chemistry*
  • Flow Cytometry / methods*
  • Humans
  • Leukocytes / immunology
  • Phycocyanin / chemistry*
  • Phycoerythrin / chemistry
  • T-Lymphocytes / immunology


  • ALLO 7
  • Antibodies, Monoclonal
  • Carbocyanines
  • Coloring Agents
  • Phycocyanin
  • Phycoerythrin