BMS166

antibody from Invitrogen Antibodies

Targeting: CTSL CTSL1, FLJ31037

Western blot (Supportive data in Antibodypedia)

ELISA (Recommended by provider)

Other assay (Supportive data in Antibodypedia)

Antibody Data

Product number

BMS166

Provider

Invitrogen Antibodies

Product name

Cathepsin L Monoclonal Antibody (33/1), eBioscience™

Antibody type

Monoclonal

Antigen

Other

Description

Description: Cathepsin L is a lysosomal cysteine proteinase consisting of a heavy chain of about 25 kDa and a light chain of about 5 kDa derived proteolytically from the same precursor. Cathepsin L has been shown to be produced by many different cell types. Since cathepsin L is capable of degrading protein constituents of the extracellular matrix it is assumed to play a crucial role in tumor progression and metastasis and a number of other disorders where the destruction of the extracellular matrix is the major cause of disease (e.g. rheumatoid arthritis, neurodegeneration). Applications Tested: ELISA, Western Blotting. Purity: >95%.

Reactivity

Human

Host

Mouse

Isotype

IgG

Antibody clone number

33/1

Vial size

100 µg

Concentration

0.1 mg/mL

Storage

4° C

References

Submitted references

SARS-CoV-2 Permissive glioblastoma cell line for high throughput antiviral screening.

Vanhulle E, Stroobants J, Provinciael B, Camps A, Noppen S, Maes P, Vermeire K
Antiviral research 2022 Jul;203:105342

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockdown of Cathepsin L was achieved by transfecting A549 with Cathepsin L specific siRNAs (Silencer® select Product # S223364, S3754). Western blot analysis (Fig. a) was performed using Whole cell extracts from the Cathepsin L knockdown cells (lane 3), non-targeting scrambled siRNA transfected cells (lane 2) and untransfected cells (lane 1). The blot was probed with Cathepsin L Monoclonal Antibody (33/1), eBioscience™ (Product # BMS166, 1 µg/mL) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:4000 dilution). Densitometric analysis of all three expected bands of this western blot is shown in histogram (Fig. b). Decrease in signal upon siRNA mediated knock down confirms that antibody is specific to Cathepsin L.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Knockout of Cathepsin L was achieved by CRISPR-Cas9 genome editing using LentiArray™ Lentiviral sgRNA (Product # A32042, Assay ID CRISPR1042354_LV) and LentiArray Cas9 Lentivirus (Product # A32064). Western blot analysis of Cathepsin L was performed by loading 30 µg of A549 Cas9 (Lane 1), andA549 Cathepsin L KO (Lane 2) whole cell extracts. The samples were electrophoresed using NuPAGE™ Novex™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with Anti-Cathepsin L Monoclonal Antibody (33/1), eBioscience™ (Product # BMS166, 1 µg/mL dilution) and Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177, 1:5,000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005). Loss of signal upon CRISPR mediated knockout (KO) using the LentiArray™ CRISPR product line confirms that antibody is specific to Cathepsin L.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Western blot was performed using Anti-Cathepsin L Monoclonal Antibody (33/1), eBioscience™(Product # BMS166) and a 25kDa band corresponding to mature form of Cathepsin L was observed across cell lines tested. An additional faint bands around 41kDa which corresponds to the Procathepsin L and 31kDa which also corresponds to the mature form of Cathepsin L was observed in A549. Whole cell extracts (30 µg lysate) of A549 (Lane 1), Hep G2 (Lane 2) and MCF 10A (Lane 3) were electrophoresed using NuPAGE™ 4-12% Bis-Tris Protein Gel (Product # NP0322BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1 ug/ml) and detected by chemiluminescence with Goat anti-Mouse IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A28177,1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).

Other assay

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Fig. 1 U87 cells have no detectable endogenous ACE2 expression but high levels of Cathepsin B. A) Different cell lines were analyzed for ACE2 expression by immunoblotting, with beta-actin as loading control. B) Same as in (A) but with the stably ACE2 transduced U87 cells, tested at an early (# 1) or late (# 32) passage of the cells. The transduced U87 cells stably express ACE2 at a very high level as compared to endogenous ACE2 in the Vero E6 cells. The same immunoblot is used for panel A and B, but with a longer exposure time for panel A in order to visualize the faint band in the Calu-3 sample. C) Comparative qPCR analysis of TMPRSS2 mRNA levels in different cells. Graph shows individual copy numbers/mul of 3 technical replicates (n = 3) as calculated from a TMPRSS2 standard. The TMPRSS2 level in Vero E6 cells was below detection limit. D) Different cell lines were analyzed for Cathepsin L (CTSL; left) and Cathepsin B (CTSB; right) expression by immunoblotting, with clathrin as loading control. For CTSL, different species are detected (indicated by the bracket): Pro-CTSL (41 kDa), glycosylated mature CTSL (31 kDa) and non-glycosylated mature CTSL (25 kDa), whereas only the Pro-CTSB form (42 kDa) is visualized. M: molecular marker in kDa, UD: undetectable. Fig. 1