Sell | L-selectin/CD62L Rabbit

Tissue/cell: rat colon tissue; 4% P‌‌araformaldehyde-fixed and paraffin-embedded;
_x000D_ Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
_x000D_ Incubation: Anti-L-Selectin Polyclonal Antibody, Unconjugated(bs-1036R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
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Tissue/cell: rat colitis tissue;4% P‌‌araformaldehyde-fixed and paraffin-embedded;
_x000D_ Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
_x000D_ Incubation: Anti-L-Selectin Polyclonal Antibody, Unconjugated(bs-1036R) 1:200, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, Cy3 conjugated(bs-0295G-Cy3)used at 1:200 dilution for 40 minutes at 37°C. DAPI(5ug/ml,blue,C-0033) was used to stain the cell nuclei
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Tissue/cell: mouse kidney tissue; 4% P‌‌araformaldehyde-fixed and paraffin-embedded;
_x000D_ Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min;
_x000D_ Incubation: Anti-L-Selectin Polyclonal Antibody, Unconjugated(bs-1036R) 1:200, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining
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Blank control: mouse spleen cells (blue).
Primary Antibody: Rabbit Anti- CD62L antibody(bs-1036R), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA;
Isotype Control Antibody: Rabbit IgG(orange) ,used under the same conditions );
Secondary Antibody: Goat anti-rabbit IgG-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA.
_x000D_ _x000D_ Protocol
_x000D_ The cells were fixed with 2% P‌‌araformaldehyde (10 min). Primary antibody (bs-1036R, 1μg /1x10^6 cells) were incubated for 30 min on the ice, followed by 1 X PBS containing 0.5% BSA + 1 0% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed._x000D_

Sample:
_x000D_ Lymph node (Mouse) Lysate at 40 ug
_x000D_ Lymph node (Rat) Lysate at 40 ug
_x000D_ Primary: Anti- CD62L (bs-1036R) at 1/300 dilution
_x000D_ Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
_x000D_ Predicted band size: 37 kD
_x000D_ Observed band size: 43 kD
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