Monoclonal antibodies (mAb) against mouse tetraspanins were CD9 (Cat. No. MCA2749, clone MF1 Bio-Rad), CD63 (Cat. No. 143902, clone NVG-2, Biolegend), CD81 (Cat. No. MCA1846, clone Eat2 Bio-Rad), and matching isotype controls rat IgG2b (Bio-Rad), rat IgG2aκ (Biolegend) and hamster IgG1 (Bio-Rad), respectively. MAb to other cell surface molecules were CD36 (Cat. No. 102602, clone HM36), CD44 (Cat. No. 103002, clone IM7) CD47 (Cat. No. 127502, clone miap301) CD98 (Cat. No. 128202, clone RL388), CD172a (Cat. No. 144002, clone P84) (all from Biolegend), and DC-STAMP (Cat. No. MABF39, clone 1A2 Millipore). Isotype controls were Armenian hamster IgG, rat IgG2bκ, rat IgG2aκ, rat IgG2aκ, rat IgG1κ, (Biolegend), and IgG2aκ (Millipore), respectively. The secondary antibodies used for flow cytometry were anti-rat IgG-FITC (Cat. No. F-9387, Sigma) and anti-hamster IgG-FITC (Cat. No. MCA2357, Bio-Rad). All antibodies were used at saturating binding concentrations.
Rat igg2b
Manufactured by Bio-Rad
Rat IgG2b is an immunoglobulin G (IgG) antibody subclass found in rats. It is a laboratory reagent commonly used in immunological research and assays.
Automatically generated - may contain errors
2 protocols using rat igg2b
1
Monoclonal Antibodies for Mouse Tetraspanins
2
Flow Cytometric Characterization of Adipose-Derived Mesenchymal Stem Cells
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The cAd-MSCs phenotypes were evaluated by performing flow cytometry. The cells were pelleted at 1500 rpm for 5 min, washed with MACS buffer (Miltenyi Biotec), and then distributed into groups according to requirement. Here we had four markers including CD34(R-Phycoerythrin conjugated, clone 1H6; BD), CD44(ALEXA FLUOR® 488-conjugated, clone YKIX337.8.7; Bio-Rad), CD45(R-Phycoerythrin conjugated clone YKIX337.8.7; Bio-Rad), and CD90(clone YKIX337.8.7; Bio-Rad). Apart from CD90, other groups were immunostained for 30 min at 4°C in the dark. CD90 requires another 30 min of incubation for secondary antibody (R-Phycoerythrin conjugated, Rabbit anti-rat, Bio-Rad). Furthermore, there were three relative isotype control antibodies that need to be prepared, including Rat igG2a (ALEXA FLUOR® 488 isotype control, Bio-Rad), Rat igG2b (R-Phycoerythrin isotype control, Bio-Rad), and Mouse igG1 (R-Phycoerythrin isotype control, Beckman Coulter), which were also incubated 30 min at 4°C in the dark. After incubation, the cells were washed and resuspended with MACS buffer. The acquisition was performed by using a FACScan (BECTON DICKINSON) with 5,000 events per sample. Lastly, data were analyzed by using FlowJo software (Tree Star).
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