The Human 1α,25-Dihydroxyvitamin D3 (1,25-(OH)₂D₃) ELISA Kit provides accurate quantitative measurement of calcitriol, the biologically active form of vitamin D that acts as a pleiotropic steroid hormone. As the principal regulator of calcium and phosphate homeostasis, this kit enables mechanistic studies of intestinal calcium absorption, bone mineralization, and parathyroid hormone (PTH) feedback, making it essential for research on osteoporosis, rickets, and osteomalacia. Because the kidney is the primary site of 1α‑hydroxylation, the assay is widely used in chronic kidney disease (CKD) and renal failure models to evaluate impaired calcitriol synthesis and its role in secondary hyperparathyroidism and mineral bone disorder. The kit further supports immunological investigations by quantifying the active vitamin D that suppresses T‑cell proliferation and modulates macrophage function, helping to dissect its protective roles in autoimmune conditions such as multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematosus, as well as in infectious disease. In oncology, the 1,25‑(OH)₂D₃ ELISA is applied to explore the link between vitamin D deficiency, disrupted signaling, and tumor progression in colorectal, breast, and prostate cancers, allowing researchers to assess how calcitriol affects cancer cell proliferation, apoptosis, and metastasis. Finally, in drug development and nutritional intervention studies, the kit serves as a critical tool to measure target engagement and systemic exposure of vitamin D receptor (VDR) agonists or antagonists, and to verify that vitamin D supplementation or novel therapeutics achieve desired active vitamin D levels in preclinical models and clinical samples.
Key Features & Benefits
- Enhanced Sensitivity with Superior Detection: Utilizes >95% pure biotinylated recombinant monoclonal antibodies paired with streptavidin-HRP conjugate (activity ≥500 U/mg) for amplified signal and a low detection limit of <0.01 ng/mL.
- High Specificity for Active Metabolite: Accurately measures 1α,25-(OH)2D3 with minimal cross-reactivity to other Vitamin D metabolites (e.g., 25-OH-D2/D3), ensuring reliable data.
- Optimized & Streamlined Protocol: A user-friendly one-step assay reduces hands-on time and simplifies the workflow for efficient processing of multiple samples.
- Exceptional Value Proposition: Obtain premium performance for less. Our Human 1α,25-(OH)2D3 ELISA Kit for sale comes with wholesale price under $100.
- Comprehensive Research Support: We offer custom ELISA kit development for unique project needs and a complimentary sample testing service to verify assay performance with your specific samples.
Enhanced Assay Principle: Biotin-Streptavidin Amplified ELISA
This kit utilizes a sensitive one-step sandwich enzyme-linked immunosorbent assay (ELISA) with signal amplification:
- Capture: A specific antibody for 1α,25-(OH)2D3 is pre-coated onto the microplate wells.
- Formation of Sandwich Complex: Samples or standards are incubated simultaneously with a biotinylated detection antibody. The target 1α,25-(OH)2D3 is bound by both antibodies.
- Signal Amplification: After washing, a Streptavidin-Horseradish Peroxidase (SA-HRP) conjugate is added, which binds with high affinity to the biotin on the detection antibody, creating an amplified detection complex.
- Color Development & Measurement: Following another wash, TMB substrate is added. The HRP catalyzes TMB’s conversion to a blue product, which turns yellow upon acid addition. The absorbance at 450 nm is measured. The signal intensity is directly proportional to the concentration of 1α,25-(OH)2D3 in the sample.
Detailed Kit Specifications & Contents
Performance Characteristics
| Parameter | Specification |
|---|---|
| Accuracy (Linearity) | Correlation coefficient R ≥ 0.9900 |
| Sensitivity | < 0.01 ng/mL |
| Detection Range | 0.1 – 2 ng/mL |
| Specificity | No significant cross-reactivity with analogous molecules |
| Precision (Repeatability) | Intra- and Inter-assay CV < 15% |
| Storage | 2-8°C, protect from light and moisture |
| Shelf Life | 6 months |
Standard Curve Concentrations
S0 to S5: 0, 0.125, 0.25, 0.5, 1, 2 ng/mL
Complete Kit Components (96T / 48T)
- Pre-coated Microplate (8 strips / 4 strips)
- Standards (0.3mL x 6 tubes)
- Sample Diluent (6mL / 3mL)
- Biotinylated Detection Antibody (10mL / 5mL)
- Streptavidin-HRP Conjugate (10mL / 5mL)
- 20x Wash Buffer (25mL / 15mL)
- Substrate A & B (6mL each / 3mL each)
- Stop Solution (6mL / 3mL)
- Plate Sealers, Comprehensive Manual, Storage Bag
Sample Handling & Preparation Guidelines
- Serum/Plasma: Collect using sterile, pyrogen-free tubes. Centrifuge at 3000×g for 10-30 minutes to obtain a clear supernatant free of cells or particles.
- Cell Culture Supernatant & Tissue Homogenate: Clarify by centrifugation at 3000×g for 10 minutes.
- Storage: For long-term storage, aliquot samples and freeze at -20°C. Avoid repeated freeze-thaw cycles. Thaw completely at room temperature and mix well before assay.
Essential Workflow & Notes
- Preparation: Equilibrate all kit components to room temperature for 20 minutes. Dilute the 20x Wash Buffer 1:20 with distilled water.
- Sample & Standard Addition: Add 50µL of standards or pre-diluted samples (10µL sample + 40µL diluent) to appropriate wells. The S0 standard serves as the blank/zero control.
- Incubation with Detection Reagents: Add 100µL of the prepared detection antibody/SA-HRP mixture to each well (except blank). Incubate at 37°C for 60 minutes.
- Washing: Wash the plate 5 times thoroughly using the prepared wash buffer.
- Substrate Reaction: Add 50µL each of Substrate A and B. Incubate at 37°C for 15 minutes in the dark.
- Stop & Read: Add 50µL Stop Solution. Measure the absorbance at 450 nm within 15 minutes.
Note: Final sample concentration = Calculated concentration from curve × dilution factor (5).
Materials Required But Not Supplied
- Microplate Reader (450 nm filter)
- Precision Pipettes and Disposable Tips
- 37°C Incubator or Water Bath
- Deionized or Distilled Water
Primary Research Applications
- Bone & Mineral Metabolism Research: Investigate the role of active Vitamin D in calcium regulation, osteoporosis, and renal osteodystrophy.
- Immunology & Autoimmunity: Study the immunomodulatory effects of 1α,25-(OH)2D3 in diseases like multiple sclerosis, rheumatoid arthritis, and immune cell differentiation.
- Cancer Research: Explore correlations between Vitamin D status, receptor expression, and outcomes in various cancers.
- Endocrine & Nutritional Studies: Assess Vitamin D sufficiency, metabolism, and its impact on overall health.
Related Products (Recommended for Cytokine Studies)
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- 1-3-β-D-glucan QuickDetect (Human) ELISA Kit – For Invasive Fungal Biomarker Detection
- Human 2′,3′-cGAMP ELISA Kit – Precision Tool for Innate Immunity & Oncology Research
Ordering, Customization & Value-Added Services
Purchase the Standard Kit: Buy the Human 1α,25-(OH)2D3 ELISA Kit online from Yanda Bio. Contact us for a specific quote, with bulk order pricing maintained under $100.
Custom ELISA Development Service: Require a modified assay range, different sample type validation, or unique conjugate? Our R&D team can develop a tailored solution.
Free Sample Testing Service: Validate the kit’s applicability for your project risk-free. Send us your samples; we will run the assay and provide a complete data report.
Choose Yanda Bio for reliable tools and expert support. Contact us today to order, request a custom quote, or schedule your complimentary sample analysis.
Frequently Asked Questions (FAQ)
Q: What is the advantage of the biotin-streptavidin detection system?
A: This system provides significant signal amplification. Multiple biotin molecules on the detection antibody bind multiple streptavidin-HRP complexes, greatly enhancing sensitivity and allowing detection of very low analyte levels (<0.01 ng/mL).
Q: Can this kit distinguish between 1α,25-(OH)2D2 and D3?
A: This kit is specifically designed for the D3 form (cholecalciferol-derived). Please check the cross-reactivity data in the manual or contact us for specifics regarding D2.
Q: How do you offer such competitive pricing?
A: Our optimized manufacturing and direct-to-researcher sales model eliminate intermediary costs, allowing us to offer high-performance Human 1α,25-(OH)2D3 ELISA Kits for sale at wholesale prices under $100.
Q: Is this kit suitable for clinical diagnosis?
A: No. This product is for Research Use Only (RUO). It is not intended for use in diagnostic procedures.
Disclaimer
- This kit is designed for research purposes only. It is not validated or approved for use in clinical, diagnostic, or therapeutic procedures involving humans.
- Users must adhere strictly to the provided instructions. The manufacturer is not liable for consequences arising from the misuse or modification of the prescribed protocol.
