Human Cathepsin D ELISA Kit

Catalog No.: BEK1023

Size: 96T

Range: 156 pg/ml-10,000 pg/ml

Sensitivity < 10 pg/ml

Storage and Expiration: Store at 2-8℃ for 6 months, or at -20℃ for 12 months. 

Application: For quantitative detection of Cathepsin D in human serum, plasma or cell culture supernatants.

Introduction

Cathepsin D is one of the lysosomal proteinases. It is ubiquitously expressed and is involved in proteolytic degradation, cell invasion, and apoptosis. It is an aspartic protease that depends critically on protonation of its active site Asp residue and gets activated at pH 5 in endosome of hepatocytes where it degrades insulin. Mutations in the CTSD gene are involved in the pathogenesis of several diseases, including breast cancer and possibly Alzheimer disease. It also has been used as a breast cancer tumor marker.

Principle of the Assay  

This kit was based on sandwich enzyme-linked immune-sorbent assay technology. Anti-Cathepsin D polyclonal antibody was pre-coated onto 96-well plates. And the biotin conjugated anti-Cathepsin D polyclonal antibody was used as detection antibodies. The standards, test samples and biotin conjugated detection antibody were added to the wells subsequently, and wash with wash buffer. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away with wash buffer. TMB substrates were used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the Cathepsin D amount of sample captured in plate. Read the O.D. absorbance at 450nm in a microplate reader, and then the concentration of Cathepsin D can be calculated.

Kit components

1. One 96-well plate pre-coated with anti-human Cathepsin D antibody
2. Lyophilized human Cathepsin D standards: 2 tubes (10 ng / tube)                     
3. Sample / Standard diluent buffer: 30ml
4. Biotin conjugated anti-human Cathepsin D antibody (Concentrated): 130μl. Dilution: 1:100
5. Antibody diluent buffer: 12ml
6. Avidin-Biotin-Peroxidase Complex (ABC) (Concentrated): 130μl. Dilution: 1:100
7. ABC diluent buffer: 12ml
8. TMB substrate: 10ml
9. Stop solution: 10ml
10. Wash buffer(25X): 30ml

Note: Reconstitute standards and test samples with Kit Component 3.

Material Required But Not Provided

1. 37℃ incubator
2. Microplate reader (wavelength: 450nm)
3. Precise pipette and disposable pipette tips
4. Automated plate washer
5. ELISA shaker
6. 1.5ml of Eppendorf tubes
7. Plate cover
8. Absorbent filter papers
9. Plastic or glass container with volume of above 1L