Anti-Cystatin SN/CST1 antibody [EPR25933-111]

• IP

Supplier Data

Immunoprecipitation - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (AB307416)

Cystatin SN/CST1 was immunoprecipitated from 0.35 mg SW480 whole cell lysate with ab307416 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab307416 at 1/1000 dilution. VeriBlot for IP secondary antibody(HRP)(ab131366) was used at 1/5000 dilution. Lane 1 : SW480 (human colorectal adenocarcinoma epithelial cell) whole cell lysate Lane 2 : ab307416 IP in SW480 whole cell lysate Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab307416 in SW480 whole cell lysate Blocking and dilution buffer and concentration : 5% NFDM/TBST. Exposure time : 15 seconds

All lanes:

Immunoprecipitation - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (ab307416) at 1/30 dilution

Secondary

All lanes:

Immunoprecipitation - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/5000 dilution

false

Exposure time: 15s

• WB

Lab

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (AB307416)

Western blot : Rabbit Monoclonal[EPR25933-111] to Cystatin SN/CST1 ab307416 staining at 1/1000 dilution; Mouse anti GAPDH ab8245 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 14 kDa. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc Milk in TBS-0.1 % Tween 20 (TBS-T) before incubation with primary antibodies overnight at 4 C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit HRP (H+L) and Goat anti-Mouse 680RD at 1/20,000 dilution. This blot was developed using a high-sensitivity ECL substrate, allowing for the detection of proteins in the mid-femtogram range.

All lanes:

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (ab307416) at 1/1000 dilution

Lane 1:

Wild-type A549 BFA (5g/mL,6h) at 20 µg

Lane 2:

Wild-type A549 BFA (0 g/mL,6h) at 20 µg

Lane 3:

CST1 knockout A549 BFA (5 g/mL, 6 h) at 20 µg

Lane 4:

CST1 knockout A549 BFA (0 g/mL, 6 h) at 20 µg

Lane 5:

SW480 at 20 µg

Lane 6:

HCT 116 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit HRP (H+L) & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 14 kDa

Observed band size: 14 kDa

true

Exposure time: 5s

• WB

Supplier Data

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (AB307416)

Blocking and diluting buffer and concentration : 5% NFDM/TBST Low expression : HCT116 (PMID : 24357805, PMID : 28300829, PMID : 19513549), 293T (PMID : 24357805). In Western blot, anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. Anti- Bovine Serum Albumin antibody (ab192603) loading control staining at 1/1000 dilution. Exposure time : 48 aeconds

All lanes:

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (ab307416) at 1/1000 dilution

Lane 1:

MDA-MB-231 (human breast adenocarcinoma epithelial cell) cytoplasmic fraction at 20 µg

Lane 2:

SW480 (human colorectal adenocarcinoma epithelial cell), whole cell lysate at 20 µg

Lane 3:

HCT116 (human colorectal carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 4:

293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

Lane 5:

MDA-MB-231 culture supernatant at 10 µL

Lane 6:

SW480 culture supernatant at 10 µL

Lane 7:

HCT116 culture supernatant at 10 µL

Lane 8:

293T culture supernatant at 10 µL

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

Observed band size: 16 kDa

false

Exposure time: 48s

• WB

Supplier Data

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (AB307416)

Blocking and diluting buffer and concentration : 5% NFDM/TBST This antibody does not cross-react with human CST2 and CST4. In Western blot, anti-His antibody (ab213204) staining at 1/5000 dilution. Anti-GAPDH antibody (ab181602) loading control staining at 1/200000 dilution. Exposure time : 15 seconds

All lanes:

Western blot - Anti-Cystatin SN/CST1 antibody [EPR25933-111] (ab307416) at 1/1000 dilution

Lane 1:

293T cells transfected with a human CST1 expression vector containi a myc-his tag whole cell lysate at 20 µg

Lane 2:

293T cells transfected with a human CST2 expression vector containi a myc-his tag whole cell lysate at 20 µg

Lane 3:

293T cells transfected with a human CST4 expression vector containi a myc-his tag whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Observed band size: 16 kDa

false

Exposure time: 15s