Recombinant Human Carbonic Anhydrase 9 (CA9) (carrier-free)

Product Details

Source

Recombinant human CA9, amino acid Pro59-Asp414 (Accession No. Q16790) was expressed in CHO cells.

Molecular Mass

The 365 amino acid recombinant protein has a predicted molecular mass of approximately 42.5 kD. The DTT-reduced protein migrates at approximately 50 kD and non-reduced protein migrates at approximately 100 kD by SDS-PAGE.

Purity

> 95%, as determined by Coomassie stained SDS-PAGE.

Formulation

0.22 µm filtered protein solution is in 25mM Tris, 150mM NaCl, pH 7.5

Endotoxin Level

Less than 0.1 EU per µg cytokine as determined by the LAL method.

Concentration

25 µg size is bottled at 200 µg/mL. 100 µg size and larger sizes are lot-specific and bottled at the concentration indicated on the vial. To obtain lot-specific concentration and expiration, please enter the lot number in our Certificate of Analysis online tool.

Storage & Handling

Unopened vial can be stored between 2°C and 8°C for up to 2 weeks, at -20°C for up to six months, or at -70°C or colder until the expiration date. For maximum results, quick spin vial prior to opening. The protein can be aliquoted and stored at -20°C or colder. Stock solutions can also be prepared at 50 - 100 µg/mL in appropriate sterile buffer, carrier protein such as 0.2 - 1% BSA or HSA can be added when preparing the stock solution. Aliquots can be stored between 2°C and 8°C for up to one week and stored at -20°C or colder for up to 3 months. Avoid repeated freeze/thaw cycles.

Activity

The esterase activity of CA9 is used to measure its specific activity (SA). The SA is > 20 pmole/min/µg, at an absorbance of 400 nm with 2 mM of enzymatic substrate (4-Nitrophenyl acetate) and 2 µg enzyme in 50 µL of 12.5 mM Tris, 75 mM NaCl, pH 7.5.

Application

Bioassay

Application Notes

The esterase activity of Human Carbonic Anhydrase 9 (hCA9) activity is used to measure its specific activity (SA). The SA is > 20 pmole/min/μg, at an absorbance of 400 nm with 2 mM of enzymatic substrate (4-Nitrophenyl acetate) and 2 μg enzyme in 50 μL of 12.5 mM Tris, 75 mM NaCl, pH 7.5. The increase of the product is monitored by increase in intensity of fluorescence at 400 nm.

 

Materials

1. Assay Buffer: 12.5 mM Tris, 75 mM NaCl, pH 7.5

2. Recombinant Human Carbonic Anhydrase 9

3. 2 mM of enzymatic substrate (4-Nitrophenyl acetate) and 2 μg enzyme in 50 μL of 12.5 mM Tris, 75 mM NaCl, pH 7.5. 

4. 4-Nitrophenol

 

Activity assay procedures

1. Prepare hCA9 to 40 μg/mL in Assay Buffer

2. Dilute 4-Nitrophenol to 0.4 mM in assay buffer. 

3. Add 100 µL of the 0.4 mM 4-Nitrophenol or 40 μg/mL hCA9 into the top concentration well

4. In a 96-well black/clear bottom plate, prepare in 1:2 serial dilutions of 4-Nitrophenol or hCA9 in assay buffer, 50 μL/well, with the top concentration of 0.4 mM (2X) 4-Nitrophenol or 40 μg/ml (2X) hCA9 Dilute substrate to 2 mM in Assay Buffer

5. Read the product formation by measuring at 400 nm in kinetic mode for 5 minutes.

6. Calculate specific activity using the equation below: 

 

* Adjusted for substrate blank

**Derived using calibration standard 4-Nitrophenol  

 

Per well

Recombinant human CA9: 1 µg

Substrate: 1 mM 

BioLegend carrier-free recombinant proteins provided in liquid format are shipped on blue-ice. Our comparison testing data indicates that when handled and stored as recommended, the liquid format has equal or better stability and shelf-life compared to commercially available lyophilized proteins after reconstitution. Our liquid proteins are validated in-house to maintain activity after shipping on blue ice and are backed by our 100% satisfaction guarantee. If you have any concerns, contact us at tech@biolegend.com.

Antigen Details

Structure

Monomor

Distribution

A431 human epithelial carcinoma cells line, U87 MG human glioblastoma/astrocytoma cell line, cancer cells

Function

CA9 interconverts carbon dioxide and bicarbonate to maintain acid-base balance in blood and other tissues, and to help transport carbon dioxide out of tissues. HIV-1 infection (VSV-G pseudotyped) of CEMT4 T cells down-regulates plasma membrane expression of CA9.

Ligand/Receptor

CO₂ and H₂O

Bioactivity

The esterase activity of CA9 is used to measure its specific activity.

Cell Type

Basophils, Mast cells, Monocytes, Neutrophils

Biology Area

Angiogenesis, Apoptosis/Tumor Suppressors/Cell Death, Cell Motility/Cytoskeleton/Structure, Cell Proliferation and Viability, Immunology

Molecular Family

CD Molecules

Antigen References

1. Uda NR, et al. 2015. J Enzyme inhib Med Chem. 6:955-60.
2. Pastorekova S, et al. 2019. Cancer Metastasis Rev. 38:65-77.
3. Pastorek J, et al. 1994. Oncogene. 9:2877-88.
4. Loncaster JA, et al. 2001. Cancer Res. 61:6394-9.
5. Kaluzova, M, et al. 2004. Mol Cell Biol. 24:5757-66.
6. Pan P, et al. 2006. J Physiol. 571:319-27.
7. Zhou GX, et al. 2010. Urology. 75:257-61.

Gene ID

768 View all products for this Gene ID

UniProt

View information about Carbonic Anhydrase 9 on UniProt.org

Documentation

• Technical Data Sheet (pdf)
• Certificate of Analysis
• Safety Data Sheet

Reviews

Related Pages & Pathways

Related FAQs

Why choose BioLegend recombinant proteins?

     • Each lot of product is quality-tested for bioactivity as indicated on the data sheet.
     • Greater than 95% Purity or higher, tested on every lot of product.
     • 100% Satisfaction Guarantee for quality performance, stability, and consistency.
     • Ready-to-use liquid format saves time and reduces challenges associated with reconstitution.
     • Bulk and customization available. Contact us.
     • Learn more about our Recombinant Proteins.

How does the activity of your recombinant proteins compare to competitors?

We quality control each and every lot of recombinant protein. Not only do we check its bioactivity, but we also compare it against other commercially available recombinant proteins. We make sure each recombinant protein’s activity is at least as good as or better than the competition’s. In order to provide you with the best possible product, we ensure that our testing process is rigorous and thorough. If you’re curious and eager to make the switch to BioLegend recombinants, contact your sales representative today!

What is the specific activity or ED50 of my recombinant protein?

The specific activity range of the protein is indicated on the product datasheets. Because the exact activity values on a per unit basis can largely fluctuate depending on a number of factors, including the nature of the assay, cell density, age of cells/passage number, culture media used, and end user technique, the specific activity is best defined as a range and we guarantee the specific activity of all our lots will be within the range indicated on the datasheet. Please note this only applies to recombinants labeled for use in bioassays. ELISA standard recombinant proteins are not recommended for bioassay usage as they are not tested for these applications.

Have your recombinants been tested for stability?

Our testing shows that the recombinant proteins are able to withstand room temperature for a week without losing activity. In addition the recombinant proteins were also found to withstand four cycles of freeze and thaw without losing activity.

Does specific activity of a recombinant protein vary between lots?

Specific activity will vary for each lot and for the type of experiment that is done to validate it, but all passed lots will have activity within the established ED50 range for the product and we guarantee that our products will have lot-to-lot consistency. Please conduct an experiment-specific validation to find the optimal ED50 for your system.

How do you convert activity as an ED50 in ng/ml to a specific activity in Units/mg?

Use formula Specific activity (Units/mg) = 10^6/ ED50 (ng/mL)

Certificate of Analysis