Antibody Type: Rabbit Polyclonal

Application: WB,IHC-P,IHC-Fr,IF,FCM

Reactivity: Human,Mouse,Rat (predicted:Dog,Pig,Cow,Rabbit)

IgG

Human,Mouse,Rat (predicted:Dog,Pig,Cow,Rabbit)

1. Paraformaldehyde-fixed, paraffin embedded (Rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15 min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 min; Blocking buffer (normal goat serum) at 37°C for 30 min; Antibody incubation with (Acid sphingomyelinase) Polyclonal Antibody, Unconjugated (TMAB-00037) at 1:400 overnight at 4°C, followed by operating according to SP Kit (Rabbit) instructionsand DAB staining.
2. Sample:
HepG2 (human) cell Lysate at 30 μg
MCF-7 (human) cell Lysate at 30 μg
A431 (human) cell Lysate at 30 μg
Hale (human) cell Lysate at 30 μg
Primary: Anti-Acid sphingomyelinase (TMAB-00037) at 1/300 dilution
Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution
Predicted band size: 64 kDa
Observed band size: 69 kDa
3. Blank control: A431. Primary Antibody (green line): Rabbit Anti-Acid sphingomyelinase antibody (TMAB-00037)
Dilution: 2 μg/10^6 cells;
Isotype Control Antibody (orange line): Rabbit IgG.
Secondary Antibody: Goat anti-rabbit IgG-PE
Dilution: 1 μg/test.
Protocol
The cells were fixed with 4% PFA (10 min at room temperature) and then permeabilized with 0.1% PBST for 20 min at room temperature. The cells were then incubated in 5% BSA to block non-specific protein-protein interactions for 30 min at room temperature. Cells stained with Primary Antibody for 30 min at room temperature. The secondary antibody used for 40 min at room temperature.

WB,IHC-P,IHC-Fr,IF,FCM

WB: 1:500-2000; IHC-P: 1:100-500; IHC-Fr: 1:100-500; IF: 1:100-500; FCM: 2ug/Test

Polyclonal

KLH conjugated synthetic peptide: human Acid sphingomyelinase

Human

6609

Associated Proteins,Neurodegenerative disease,Lipid metabolism