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55279S 100 µl (50 tests) $341.00.0
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REACTIVITY SENSITIVITY MW (kDa) Isotype
H M R Endogenous Rabbit IgG
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Immunofluorescence (Immunocytochemistry)

Confocal immunofluorescent analysis of Malme-3M (left) and MCF7 (right) cells using Cathepsin (D1C7Y) (Alexa Fluor® 488 Conjugate) (green) and β-Actin (8H10D10) Mouse mAb #3700 (red). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).

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Immunofluorescence (Immunocytochemistry)

A. Solutions and Reagents

NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.

  1. 20X Phosphate Buffered Saline (PBS): (9808) To prepare 1L 1X PBS: add 50 ml 20X PBS to 950 ml dH2O, mix. Adjust pH to 8.0.
  2. Methanol, 100%
  3. Blocking Buffer (1X PBS / 5% normal goat serum (#5425) / 0.3% Triton™ X-100): To prepare 10 ml: add 0.5 ml normal goat serum and 0.5 ml 20X PBS to 9.0 ml dH2O, mix. While stirring, add 30 µl Triton™ X-100.
  4. Antibody Dilution Buffer (1X PBS / 1% BSA / 0.3% Triton X-100): To prepare 10 ml, add 30 µl Triton™ X-100 to 10 ml 1X PBS. Mix well then add 0.1 g BSA (9998), mix.
  5. Prolong® Gold AntiFade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).

B. Specimen Preparation - Cultured Cell Lines (IF-IC)

NOTE: Cells should be grown, treated, fixed and stained directly in multi-well plates, chamber slides or on coverslips.

  1. Aspirate liquid, then cover cells to a depth of 2–3 mm with ice-cold 100% methanol.
  2. Allow cells to fix for 15 minutes at -20°C.
  3. Aspirate fixative, rinse three times in 1X PBS for 5 minutes each.
  4. Proceed with Immunostaining (Section C).

C. Immunostaining

NOTE: All subsequent incubations should be carried out at room temperature unless otherwise noted in a humid light-tight box or covered dish/plate to prevent drying and fluorochrome fading.

  1. Block specimen in Blocking Buffer for 60 minutes.
  2. While blocking, prepare primary antibody by diluting as indicated on datasheet in Antibody Dilution Buffer.
  3. Aspirate blocking solution, apply diluted primary antibody.
  4. Incubate overnight at 4°C.
  5. Rinse three times in 1X PBS for 5 minutes each.
  6. Coverslip slides with Prolong® Gold Antifade Reagent (#9071), Prolong® Gold AntiFade Reagent with DAPI (#8961).
  7. For best results, examine specimens immediately using appropriate excitation wavelength. For long-term storage, store slides flat at 4°C protected from light.

posted December 2010

Protocol Id: 221

Product Usage Information

Application Dilutions
Immunofluorescence (Immunocytochemistry) 1:50

Storage: Supplied in PBS (pH 7.2), less than 0.1% sodium azide and 2 mg/ml BSA. Store at 4°C. Do not aliquot the antibody. Protect from light. Do not freeze.

Specificity / Sensitivity

Cathepsin B (D1C7Y) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate) recognizes endogenous levels of total cathepsin B protein. This antibody detects the heavy chain subunit of cathepsin B.


Species Reactivity: Human, Mouse, Rat

Source / Purification

Monoclonal antibody is produced by immunizing animals with recombinant protein specific to the heavy chain subunit of human cathepsin B protein.

Product Description

This Cell Signaling Technology antibody is conjugated to Alexa Fluor® 488 fluorescent dye and tested in-house for immunofluorescent analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated Cathepsin B (D1C7Y) XP® Rabbit mAb #31718.


Cathepsin B (CSTB), part of the papain family of proteases, is a widely expressed lysosomal cysteine endopeptidase (1,2). Cathepsin B is produced from a larger precursor form, pro-cathepsin B, which runs at approximately 44 kDa on SDS-PAGE, and is proteolytically processed and glycosylated to form a mature two-chain protein containing a heavy chain (running at 27 and 24 kDa) and a light chain (5 kDa). High levels of cathepsin B are found in macrophages and osteoclasts, as well as various types of cancer cells, including lung, colon, prostate, breast, and stomach. In addition, expression of cathepsin B has been associated with multiple sclerosis (3), rheumatoid arthritis (4), and pancreatitis (5). While generally localized to lysosomes, in cancer alterations can lead to its secretion (6). Its role in tumor progression is thought to involve promotion of basement membrane degradation, invasion and metastasis (7,8). Expression can correlate with poor prognosis for a variety of forms of cancer (9-13).


1.  Chan, S.J. et al. (1986) Proc Natl Acad Sci USA 83, 7721-5.

2.  Fong, D. et al. (1986) Proc Natl Acad Sci USA 83, 2909-13.

3.  Bever, C.T. et al. (1994) Neurology 44, 745-8.

4.  Hashimoto, Y. et al. (2001) Biochem Biophys Res Commun 283, 334-9.

5.  Halangk, W. et al. (2000) J Clin Invest 106, 773-81.

6.  Berquin, I.M. and Sloane, B.F. (1996) Adv Exp Med Biol 389, 281-94.

7.  Yan, S. et al. (1998) Biol Chem 379, 113-23.

8.  Vasiljeva, O. et al. (2006) Cancer Res 66, 5242-50.

9.  Campo, E. et al. (1994) Am J Pathol 145, 301-9.

10.  Foekens, J.A. et al. (1998) J Clin Oncol 16, 1013-21.

11.  Werle, B. et al. (1999) Br J Cancer 81, 510-9.

12.  Lah, T.T. et al. (2000) Clin Cancer Res 6, 578-84.

13.  Werle, B. et al. (2000) Cancer 89, 2282-91.


Entrez-Gene Id 1508
Swiss-Prot Acc. P07858


For Research Use Only. Not For Use In Diagnostic Procedures.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
XP is a registered trademark of Cell Signaling Technology, Inc.
Alexa Fluor is a registered trademark of Life Technologies Corporation.
DRAQ5 is a registered trademark of Biostatus Limited.
This product is provided under an intellectual property license from Life Technologies Corporation. The transfer of this product is contingent on the buyer using the purchased product solely in research, including use with HCS or other automated imaging applications but excluding use in combination with DNA microarrays. The buyer must not sell or otherwise transfer this product or its components for (a) diagnostic, therapeutic or prophylactic purposes; (b) testing, analysis or screening services, or information in return for compensation on a per-test basis; (c) manufacturing or quality assurance or quality control, or (d) resale, whether or not resold for use in research. For information on purchasing a license to this product for purposes other than as described above, contact Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, CA 92008 USA or outlicensing@lifetech.com.

55279
Cathepsin B (D1C7Y) XP® Rabbit mAb (Alexa Fluor® 488 Conjugate)