Anti-Thrombin antibody

• WB

Ap

Western blot - Anti-Thrombin antibody (AB92621)

This blot was produced using a 4-12% Bis-tris gel under the MOPS buffer system. The gel was run at 200V for 50 minutes before being transferred onto a Nitrocellulose membrane at 30V for 70 minutes. The membrane was then blocked for an hour using 2% Bovine Serum Albumin before being incubated with ab92621 overnight at 4°C. Antibody binding was detected using an anti-rabbit antibody conjugated to HRP, and visualised using ECL development solution ab133406.

All lanes:

Western blot - Anti-Thrombin antibody (ab92621) at 1 µg/mL

Lane 1:

Liver (Human) Tissue Lysate at 10 µg

Lane 2:

Liver (Mouse) Tissue Lysate at 10 µg

Lane 3:

Liver (Rat) Tissue Lysate at 10 µg

Lane 4:

Plasma (Human) Total Protein Lysate at 10 µg

Lane 5:

Plasma (Mouse) Total Protein Lysate at 10 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG H&L (HRP) at 1/50000 dilution

Predicted band size: 70 kDa

Observed band size: 100 kDa,60 kDa,78 kDa

true

Exposure time: 5s

• WB

Supplier Data

Western blot - Anti-Thrombin antibody (AB92621)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Negative control : HeLa (PMID : 1463733).

Low expression : cerebellum.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 17637839 and PMID : 21131592).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Actual observed bands 33-72kDa.

All lanes:

Western blot - Anti-Prothrombin antibody [RM1252] (<a href='/en-us/products/primary-antibodies/prothrombin-antibody-rm1252-ab322113'>ab322113</a>) at 1/1000 dilution

Lane 1:

Human plasma lysate at 4 µg

Lane 2:

Human liver tissue lysate at 20 µg

Lane 3:

Human cerebellum tissue lysate at 20 µg

Lane 4:

Human heart tissue lysate at 20 µg

Lane 5:

HepG2 (human hepatocellular carcinoma epithelial cell) whole cell lysate at 20 µg

Lane 6:

Caco-2 (human colorectal adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Lane 7:

HeLa (human cervical adenocarcinoma epithelial cell) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - 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

Observed band size: 33-72 kDa,36 kDa

false

Exposure time: 180s

• WB

Supplier Data

Western blot - Anti-Thrombin antibody (AB92621)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

Low expression : Brain

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 17637839 and PMID : 21131592).

In Western blot, Anti-GAPDH antibody [EPR16891] - Loading Control (ab181602) staining at 1/200000 dilution.

Actual observed bands 33-72kDa.

Exposure times : left panel : 10 seconds; right panel : 147 seconds.

All lanes:

Western blot - Anti-Prothrombin antibody [RM1252] (<a href='/en-us/products/primary-antibodies/prothrombin-antibody-rm1252-ab322113'>ab322113</a>) at 1/1000 dilution

Lane 1:

Mouse plasma lysate at 10 µg

Lane 2:

Mouse liver tissue lysate at 20 µg

Lane 3:

Mouse brain tissue lysate at 20 µg

Lane 4:

Mouse heart tissue lysate at 20 µg

Lane 5:

Mouse kidney tissue lysate at 20 µg

Lane 6:

Rat plasma lysate at 10 µg

Lane 7:

Rat liver tissue lysate at 20 µg

Lane 8:

Rat brain tissue lysate at 20 µg

Lane 9:

Rat heart tissue 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: 33-72 kDa,36 kDa

false

• WB

CiteAb

Western blot - Anti-Thrombin antibody (AB92621)

Thrombin western blot using anti-Thrombin antibody ab92621. Publication image and figure legend from Kobori, T., Hamasaki, S., et al., 2018, Front Immunol, PubMed 29559970.

ab92621 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab92621 please see the product overview.

Thrombin contributes to macrophage (Mφ) M2 polarization and angiogenic capacity through proteolytic modification for osteopontin (OPN). (A) The mRNA expression level of Prothrombin relative to glyceraldehyde-3-phosphate dehydrogenase (Gapdh) was analyzed by reverse transcription polymerase chain reaction in each Mφ subset and were normalized to Mφ (–), n = 7 [***p < 0.001, **p < 0.01 vs. untreated, ##p < 0.01 vs. interleukin (IL)-10 alone]. (B,C) The protein expression levels of (B) thrombin or (C) OPN N-Half relative to GAPDH were measured by western blot in each Mφ subset and were normalized to Mφ (–). Lower panels are typical images of each protein. (B)n = 8 (***p < 0.001, *p < 0.05 vs. untreated, #p < 0.05 vs. IL-10 alone). (C)n = 16 (***p < 0.001, *p < 0.05 vs. untreated, ##p < 0.01, #p < 0.05 vs. IL-10 alone, †††p < 0.001 vs. IL-10 + IL-18). (D) Representative confocal laser scanning immunofluorescence images of OPN (red), thrombin (green), and their merge with DAPI (blue) in each Mφ subset. Scale bar represents 20 μm. Higher magnification images are from the white rectangle region in merged panel of Mφ (IL-10 + IL-18). Scale bar represents 10 μm. (E) Relative mean fluorescence intensity (MFI) of CD163 was measured by FACS analysis in each Mφ subset. Hirudin, a specific thrombin inhibitor, was used at 1 μg/mL, n = 3 (***p < 0.001 vs. untreated, ###p < 0.001 vs. IL-10 alone, †††p < 0.001 vs. IL-10 + IL-18). (F) The total areas and lengths of tube-like structures were determined by the Matrigel tube formation assay where b.End5 were cocultured with each Mφ subset. Hirudin was used at 1 μg/mL, n = 6 (***p < 0.001, **p < 0.01 vs. untreated, ##p < 0.01, #p < 0.05 vs. IL-10 alone, †††p < 0.001 vs. IL-10 + IL-18). All data are expressed as means ± SEM and were analyzed by a one-way ANOVA followed by Tukey's test.

false