准备和存储
推荐的实验流程
An isotype control should be used at the same concentration as the antibody of interest (e.g., ≤ 1 µg/million cells for flow cytometry). For tandem conjugates incorporating PerCP (e.g., PerCP-Cy5.5), the excitation and emission properties of PerCP and the kinetics of energy exchange between the fluorochromes of the tandem dye may limit their effectiveness on high-speed and/or flow cytometers.
商品通知
- Since applications vary, each investigator should titrate the reagent to obtain optimal results.
- Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
- Although hamster immunoglobulin isotypes have not been well defined, BD Biosciences Pharmingen has grouped Armenian and Syrian hamster IgG monoclonal antibodies according to their reactivity with a panel of mouse anti-hamster IgG mAbs. A table of the hamster IgG groups, Reactivity of Mouse Anti-Hamster Ig mAbs, may be viewed at http://www.bdbiosciences.com/documents/hamster_chart_11x17.pdf.
- PerCP is a photosynthetic accessory pigment from Glenodinium species of dinoflagellates, which is excited by the 488-nm light of an Argon ion laser and fluoresces at 675 nm. Therefore, PerCP-labelled antibodies can be used with FITC- and R-PE–labelled reagents in most single-laser flow cytometers with no significant spectral overlap of PerCP fluorescence with that of FITC or R-PE. PerCP has been reported to undergo significant photobleaching, the magnitude of which increases as laser power is increased or beam focus is narrowed. For third-color flow¬cytometric analysis using ≥25-mW laser power, we recommend PE-Cy5-, PE-Cy7–, or PerCP-Cy5.5-conjugated reagents.
- For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
- Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
The A19-3 monoclonal antibody specifically binds to the hapten, trinitrophenol (TNP). TNP is not expressed on human, mouse, or rat cells. The immunoglobulin from clone A19-3 was selected as an isotype control following screening for low background staining on a variety of mouse and human tissues.
研发参考 (4)
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Afar B, Merrill J, Clark EA. Detection of lymphocyte subsets using three-color/single-laser flow cytometry and the fluorescent dye peridinin chlorophyll-alpha protein. J Clin Immunol. 1991; 11(5):254-261. (Methodology). 查看参考
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Greimers R, Trebak M, Moutschen M, Jacobs N, Boniver J. Improved four-color flow cytometry method using fluo-3 and triple immunofluorescence for analysis of intracellular calcium ion ([Ca2+]i) fluxes among mouse lymph node B- and T-lymphocyte subsets. Cytometry. 1996; 23(3):205-217. (Methodology). 查看参考
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Shapiro HM. Practical Flow Cytometry, 3rd Edition. New York: Wiley-Liss, Inc; 1995:280-281.
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Waggoner AS, Ernst LA, Chen CH, Rechtenwald DJ. PE-CY5. A new fluorescent antibody label for three-color flow cytometry with a single laser. Ann N Y Acad Sci. 1993; 677:185-193. (Methodology). 查看参考
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