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PerCP Conjugated to Secondary Antibodies Jackson ImmunoResearch

Nuovo fluorocromo PerCP coniugato ad anticorpi secondari.

 
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PerCP is a fluorescent peridinin-chlorophyll protein complex isolated from dinoflagellates. We offer the form found in Dinophyceae sp. with a molecular weight of about 35.5 kDa. It has a broad spectrum of excitation with a main peak at 472 nm, and a long Stokes shift to an emission peak at 677 nm (Figure 1).
     
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Figure 1. Relative shape and position of spectra in the peak region of excitation (blue) and emission (purple) for PerCP conjugated to an affinity-purified secondary antibody from Jackson ImmunoResearch. Quantitative comparisons should not be made since peak heights have been normalized. Spectra were obtained with an M-Series spectrofluorometer system from Photon Technology International, Inc.
     
PerCP conjugates of highly adsorbed secondary antibodies are offered to label unconjugated primary antibodies, and PerCP-streptavidin is offered to label biotinylated primary or secondary  antibodies  (Figures 2 and 3).  Two practical  labeling  protocols are possible with these products. Compared with a single-step PerCP-conjugated primary antibody (Figure 2), about the same level of fluorescence is obtained with a two-step procedure using a biotinylated primary antibody and PerCP-conjugated streptavidin. A consistent, slightly higher signal is achieved by using an unconjugated primary antibody and PerCP-conjugated secondary antibody. Although three-step procedures are usually undesirable for flow cytometry, a somewhat greater amplification may be obtained with unconjugated primary antibody, biotinylated secondary antibody, and PerCP-conjugated streptavidin (Figure 3).
     
percpflowa.gif   percpflowb.gif
Figure 2. Human peripheral lymphocytes were stained either directly with PerCP-conjugated anti-CD3 (orange) from BioLegend, or indirectly with biotin-conjugated anti-CD3 (B-D Pharmingen) and PerCP-streptavidin either from Jackson ImmunoResearch (blue), BioLegend (purple), or B-D Pharmingen (red). For further comparison, cells were stained with unconjugated anti-CD3 (B-D Pharmingen) and PerCP-conjugated secondary antibody from Jackson Immuno-Research (green). Stained cells were analyzed in a dual-laser FACSCalibur flow cytometer (Becton Dickinson).   Figure 3. Human peripheral lymphocytes were stained with unconjugated anti-CD3 (B-D Pharmingen) and PerCP-conju-gated secondary antibody from Jackson ImmunoResearch (green), or with biotin-conjugated anti-CD3 (B-D Pharmingen) and PerCP-streptavidin from Jackson ImmunoResearch (blue). For further comparison, cells were stained with unconjugated anti-CD3 (B-D Pharmingen), biotinylated secondary antibody and PerCP-streptavidin from Jackson ImmunoResearch (red). Stained cells were analyzed in a dual-laser FACSCalibur flow cytometer (Becton Dickinson)
     
PerCP conjugates may be used alone or with DyLight 488 (or FITC) and R-PE for one- to three-color analyses with a single-laser flow cytometer equipped with an argon laser emiting at 488 nm. Up to four-color analyses with low compensation are easily achieved by adding APC-conjugated antibodies with 633 or 635 nm excitation provided by a dual-laser flow cytometer (Figure 4).
     
 flow4colorex3.gif    flow4colorem3.gif
Figure 4. Excitation spectra (left) for PerCP-(blue), DyLight 488/FITC- (green), R-PE- (red), and APC- (brown) conjugated secondary antibodies from Jackson ImmunoResearch. Emission spectra (right) for DyLight 488/FITC- (green), R-PE- (pink), APC- (brown), and PerCP- (purple) conjugated secondary antibodies from Jackson ImmunoResearch. Quantitative comparisons should not be made since peak heights have been normalized. All spectra were obtained with an M-Series spectrofluorometer system from Photon Technology International, Inc.
 
See Table of NEW PerCP Conjugates here

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