Recombinant Human CA8 Protein: Research Guidelines & Applications

Recombinant Human CA8 Protein is a research-grade recombinant protein derived from the human carbonic anhydrase 8 gene, offering a reliable tool for investigating CA8-associated signaling pathways, protein–protein interactions, and neurological disease mechanisms in vitro and in vivo.

Product Name

Recombinant Human CA8 Protein

SKU

rp369214

Gene Target

CA8 (Carbonic Anhydrase 8)

Species Origin

Human (Homo sapiens)

Application

Biochemical assays, protein interaction studies, neuroscience research

Storage Recommendation

Avoid repeated freeze-thaw cycles; store aliquots at -80°C for long-term use

Reference Number

absea#PP-11253

Background: CA8 Gene and Protein Overview

Carbonic anhydrase 8 (CA8), also known as carbonic anhydrase-related protein I (CARP I), is a member of the carbonic anhydrase superfamily. Unlike catalytically active carbonic anhydrases, CA8 lacks key active-site residues required for zinc coordination and therefore has no enzymatic CO₂ hydration activity. Instead, CA8 functions primarily as a regulatory or scaffolding protein.

CA8 is highly expressed in the cerebellum and other regions of the central nervous system. It has been reported to interact with inositol 1,4,5-trisphosphate receptor type 1 (IP3R1), modulating intracellular calcium signaling. Mutations or dysregulation of CA8 have been associated with cerebellar ataxia and intellectual disability in human patients, underscoring its importance in neurological function.

Recombinant Human CA8 Protein (SKU: rp369214) enables researchers to investigate these biological roles under controlled in vitro conditions, serving as a reference standard or functional reagent in biochemical and cell-based assays.

Protein Properties and Quality Specifications

Recombinant Human CA8 Protein is produced using recombinant expression systems to ensure consistent batch-to-batch quality. Researchers should consult the product-specific Certificate of Analysis (CoA) for confirmed specifications including purity, molecular weight, endotoxin levels, and biological activity data, as these parameters are determined empirically for each production lot.

General considerations for recombinant CA8 protein include:

• Purity: Typically assessed by SDS-PAGE and/or HPLC; a CoA-confirmed purity value should be used for experimental planning.
• Molecular Weight: The theoretical molecular weight of full-length human CA8 is approximately 35 kDa; recombinant constructs may include tags that shift apparent molecular weight on gel.
• Formulation: Supplied in a carrier buffer; exact buffer composition and protein concentration are detailed on the product CoA.
• Endotoxin: Low-endotoxin preparations are recommended for cell-based assays; verify endotoxin levels from the CoA prior to use.

Always refer to the most current product datasheet available at Aladdin Scientific for authoritative specifications.

Storage, Handling, and Reconstitution Guidelines

Proper handling of Recombinant Human CA8 Protein is essential to maintain its integrity and functionality throughout the experiment lifecycle. Follow the guidelines below:

Storage

• Store the protein at -80°C for long-term preservation.
• For short-term use (up to 1 week), storage at -20°C is acceptable.
• Avoid repeated freeze-thaw cycles, as this can cause aggregation and loss of activity. Pre-aliquot the protein upon first thaw.
• Do not store at 4°C for extended periods.

Reconstitution (if lyophilized)

• Briefly centrifuge the vial before opening to consolidate any powder.
• Reconstitute with the recommended volume of sterile, deionized water or the buffer specified on the datasheet.
• Allow the protein to dissolve gently; do not vortex vigorously.
• Allow a brief equilibration period at room temperature after reconstitution before aliquoting.

Working Conditions

• Prepare working dilutions fresh on the day of use.
• Use low-protein-binding tubes and tips to minimize adsorptive losses.
• Include a carrier protein such as BSA (0.1–1%) in dilution buffers if required by the assay format.

Research Applications

Recombinant Human CA8 Protein is suitable for a range of in vitro and biochemical research applications, including but not limited to:

• Protein–Protein Interaction Studies: CA8 is known to interact with IP3R1; recombinant CA8 can be used in pull-down, co-immunoprecipitation validation, or surface plasmon resonance (SPR) assays to map interaction domains.
• ELISA Standard and Antibody Validation: Purified recombinant CA8 serves as a positive control or standard curve material in sandwich ELISA formats and for validating anti-CA8 antibodies in Western blot or immunoassay development.
• Functional Assays: Researchers investigating IP3R1-mediated calcium signaling regulation can use recombinant CA8 to reconstitute or perturb the CA8–IP3R1 complex in cell-free or cell-based systems.
• Structural Biology: Recombinant CA8 can provide material for crystallography or cryo-EM studies aimed at elucidating the three-dimensional structure of CA8 and its binding interfaces.
• Biomarker Research: Given the association of CA8 dysregulation with neurological disorders, recombinant CA8 may be used to develop or validate diagnostic assays for cerebellar ataxia and related conditions.

Researchers should validate the protein in their specific assay system and consult the product datasheet for any application-specific recommendations.

Quality Control and Troubleshooting

To obtain reliable and reproducible results with Recombinant Human CA8 Protein, implement the following quality control practices and troubleshooting strategies:

Quality Control Best Practices

• Always include a positive control (known CA8 sample) and negative control (buffer blank) in each experimental run.
• Confirm protein integrity by running a small aliquot on SDS-PAGE under reducing and non-reducing conditions prior to use in critical assays.
• Check the CoA for lot-specific purity, concentration, and activity data before commencing experiments.

Common Issues and Solutions

• Loss of Activity After Thawing: Ensure rapid thaw on ice or at room temperature; avoid warming at 37°C. Pre-aliquot to prevent repeated freeze-thaw.
• Unexpected Band Pattern on SDS-PAGE: Confirm whether the construct carries a purification tag; the tag adds to molecular weight. Degradation bands may indicate improper storage — discard compromised lots.
• High Background in ELISA: Titrate the protein concentration; ensure dilution buffers contain appropriate blocking agents such as BSA.
• Aggregation: If visible aggregates are present, briefly centrifuge (10,000 × g, 5 min, 4°C) and use the supernatant. Consider adding non-ionic detergent (e.g., 0.01% Tween-20) to dilution buffers.

For persistent technical issues, contact Aladdin Scientific technical support with the lot number from your CoA.

Frequently asked questions

What is CA8 protein and why is it important in neuroscience research?

CA8 (Carbonic Anhydrase 8) is a catalytically inactive member of the carbonic anhydrase family that functions as a regulatory protein rather than an enzyme. It is highly expressed in cerebellar Purkinje cells and interacts with the IP3R1 receptor to modulate intracellular calcium signaling. Loss-of-function mutations in CA8 are linked to cerebellar ataxia and intellectual disability, making it a key target in neurological disease research.

How should I store Recombinant Human CA8 Protein to maintain activity?

For long-term storage, keep the protein at -80°C in single-use aliquots to avoid repeated freeze-thaw cycles, which can cause aggregation and activity loss. For short-term use within one week, -20°C is acceptable. Avoid prolonged storage at 4°C and never freeze-thaw the stock vial more than once.

Which assay formats are compatible with Recombinant Human CA8 Protein (rp369214)?

Recombinant Human CA8 Protein (rp369214) is suitable for use in ELISA (as standard or positive control), Western blot validation of anti-CA8 antibodies, pull-down and co-immunoprecipitation assays for protein interaction studies, and as a substrate or binding partner in functional biochemical assays. Researchers should validate suitability in their specific assay system prior to use in critical experiments.

What should I do if I observe aggregation in my CA8 protein sample?

If aggregation is observed, briefly centrifuge the sample at 10,000 × g for 5 minutes at 4°C and use the clarified supernatant. Adding a low concentration of non-ionic detergent such as 0.01% Tween-20 to the dilution buffer may help reduce aggregation. Repeated freeze-thaw cycles and storage at 4°C are the most common causes of aggregation and should be strictly avoided.

Is Recombinant Human CA8 Protein suitable for use as an ELISA standard?

Yes, purified Recombinant Human CA8 Protein can be used to construct standard curves in sandwich ELISA systems designed to quantify CA8 levels in biological samples. Ensure that the recombinant form used for the standard curve is recognized by both capture and detection antibodies in your specific ELISA kit or assay setup. Confirm endotoxin levels from the CoA if using in cell-based assay validation.

References

1. Türeci Ö, et al. Carbonic anhydrase-related proteins: molecular cloning of CARP and identification of a novel subtype (CARP II). Journal of Biological Chemistry. 1998.
2. Hirota J, Furuichi T, Mikoshiba K. Inositol 1,4,5-trisphosphate receptor type 1 is a substrate for caspase-3 and is cleaved during apoptosis in a caspase-3-dependent manner. Journal of Biological Chemistry. 1999.
3. Nishimura H, et al. Mutations in CARP-related proteins linked to cerebellar ataxia with intellectual disability. (Representative reference — specific citation needs verification.)

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.