| Field | Specification |
|---|---|
| Target | |
| Alternative names | Protein disulfide-isomerase; PDI; Cellular thyroid hormone-binding protein; Prolyl 4-hydroxylase subunit beta; p55; P4HB; ERBA2L; PDI; PDIA1; PO4DB |
| Gene ID | |
| UniProt # | |
| Reactivity | |
| Applications | |
| Immunogen | Expression system for standard: NS0; Immunogen sequence: D18-L508 |
| Expression system | |
| Expression region | |
| Assay type | |
| Sample type(s) | Cell Culture Supernatant, Serum, Plasma |
| Sensitivity | |
| Detection range | |
| Assay time | |
| Storage | |
| Catalog no. (Mfr.) | |
| Main SKU |
Assay Principle
This sandwich ELISA quantifies human protein disulfide-isomerase (gene P4HB) in cell culture supernatants, serum and plasma (heparin, EDTA). Strips are pre-coated with a capture antibody. After the sample is added, a biotinylated detection antibody binds the captured protein, and an avidin–biotin–peroxidase complex supplies the enzyme. TMB develops color in proportion to the bound target; the reaction is stopped and read at 450 nm, and concentrations are read from the standard curve.
Protein disulfide isomerase (PDI, P4HB) is an abundant endoplasmic reticulum enzyme that forms, breaks and reshuffles disulfide bonds in newly made proteins and can also act as a chaperone. It is the β subunit of collagen prolyl 4-hydroxylase and of the microsomal triglyceride transfer protein, and is studied in protein folding and ER stress.
Kit Components
| Anti-Human P4HB Pre-coated 96-well strip microplate | 1 plate (96 wells) |
|---|---|
| Human P4HB Standard | 2 vials, 10 ng/tube |
| Human P4HB Biotinylated antibody (100x) | 100 µL |
| Avidin-Biotin-Peroxidase Complex (100x) | 100 µL |
| Sample Diluent | 30 mL |
| Antibody Diluent | 12 mL |
| Avidin-Biotin-Peroxidase Diluent | 12 mL |
| Color Developing Reagent (TMB) | 10 mL |
| Stop Solution | 10 mL |
| Wash Buffer (25x) | 20 mL |
| Adhesive plate sealers | 4 |
Also required (not supplied): microplate reader for 450 nm; incubator; automated plate washer (optional); precision pipettes for 0.5 µL through 1 mL volumes of aqueous solutions; multichannel pipettes (recommended for many samples); deionized or distilled water; 500 mL graduated cylinders; test tubes for dilution.
Kit components are not sold separately.
Performance Data
| Species | Human |
|---|---|
| Sample types | Cell culture supernatants, serum and plasma (heparin, EDTA) |
| Assay range | 93.8 pg/mL–6,000 pg/mL |
| Sensitivity | <15 pg/mL |
| Standard | Recombinant human P4HB (Asp18–Leu508), expressed in NS0 |
| Cross-reactivity | None detected against related proteins in supplier testing |
| Intra-assay CV | 4.4–7.1% (3 samples, n = 16 each) |
| Inter-assay CV | 5.7–7.7% (3 samples, n = 24 each) |
| Format | 96 wells, removable strips |
Sample dilution: start at 1:5. In the supplier’s tests, serum and plasma samples measured 2000–4000 pg/mL.
Example standard curve (OD450, pg/mL): 0 = 0.025; 93.8 = 0.073; 187.5 = 0.122; 375 = 0.22; 750 = 0.388; 1500 = 0.777; 3000 = 1.315; 6000 = 2.003. Run a new standard curve on every plate.
Storage & Handling
Store at 4 °C for up to 6 months or −20 °C for up to 12 months. Avoid repeated freeze–thaw cycles. Ships on gel ice and tolerates up to 3 days at room temperature; freeze on arrival.
Safety
Sample data
| Concentration (pg/ml) | 0 | 93.8 | 187.5 | 375 | 750 | 1500 | 3000 | 6000 |
| O.D. | 0.025 | 0.16 | 0.263 | 0.491 | 0.852 | 1.29 | 1.878 | 2.305 |
Intra/inter assay consistency
| Intra-Assay Precision | Inter-Assay Precision | |||||
|---|---|---|---|---|---|---|
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 16 | 16 | 16 | 24 | 24 | 24 |
| Mean (pg/ml) | 155 | 785 | 2454 | 177 | 790 | 2763 |
| Standard deviation | 8.84 | 55.74 | 107.98 | 10.09 | 48.19 | 212.75 |
| CV (%) | 5.7% | 7.1% | 4.4% | 5.7% | 6.1% | 7.7% |
Kit components
Description|Quantity Pre-coated 96-well strip microplate|1 Standard|2 vials Biotinylated antibody (100x)|100ul Avidin-Biotin-Peroxidase Complex (100x)|100ul Sample Diluent|30ml Antibody Diluent|12ml Avidin-Biotin-Peroxidase Diluent|12ml Color Developing Reagent (TMB)|10ml Stop Solution|10ml Wash Buffer (25x)|20ml Adhesive plate sealers|4Materials required but not provided
- Microplate Reader capable of reading absorbance at 450nm.
- Incubator.
- Automated plate washer (optional).
- Pipettes and pipette tips capable of precisely dispensing 0.5 µl through 1 ml volumes of aqueous solutions.
- Multichannel pipettes are recommended for large amount of samples.
- Deionized or distilled water.
- 500ml graduated cylinders.
- Test tubes for dilution.
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