| Field | Specification |
|---|---|
| Target | |
| Alternative names | Peptidyl-glycine alpha-amidating monooxygenase; PAM; Peptidylglycine alpha-hydroxylating monooxygenase; PHM; Peptidyl-alpha-hydroxyglycine alpha-amidating lyase; Peptidylamidoglycolate lyase; PAL; PAM |
| Gene ID | |
| UniProt # | |
| Reactivity | |
| Applications | |
| Immunogen | Expression system for standard: NS0; Immunogen sequence: M1-V817 |
| 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 EZ-Set™ kit supplies a matched capture and detection antibody pair and a standard for building your own sandwich ELISA for human peptidyl-glycine alpha-amidating monooxygenase (gene PAM) in cell culture supernatants, serum and plasma (heparin, EDTA, citrate). You coat the plate with the capture antibody, add samples and standards, detect with the biotinylated antibody and an avidin–biotin–peroxidase complex, develop with TMB and read at 450 nm. Plates, diluents, wash buffer, TMB and stop solution are not included.
Peptidylglycine α-amidating monooxygenase (PAM) is a bifunctional enzyme that adds the C-terminal amide to many peptide hormones and neuropeptides. Two domains act in sequence, a copper-dependent monooxygenase and a zinc-dependent lyase, and amidation is often needed for full peptide activity.
Kit Components
| Mouse anti-human PAM monoclonal antibody (Capture Antibody) | 500 µL, 4 µg/mL (recommended dilution 1:100) |
|---|---|
| Biotinylated goat anti-human PAM polyclonal antibody (Detection Antibody) | 500 µL (recommended dilution 1:100) |
| Avidin-Biotin-Peroxidase Complex (ABC) | 500 µL (recommended dilution 1:100) |
| Lyophilized recombinant human PAM standard | 3 × 10 ng/tube |
Performance Data
| Species | Human |
|---|---|
| Sample types | Cell culture supernatants, serum and plasma (heparin, EDTA, citrate) |
| Assay range | 156 pg/mL–10,000 pg/mL |
| Standard | Recombinant human PAM (Met1–Val817), expressed in NS0 |
| Cross-reactivity | None detected against related proteins in supplier testing |
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 | 156 | 312 | 625 | 1,250 | 2,500 | 5,000 | 10,000 |
| O.D. | 0.028 | 0.054 | 0.161 | 0.387 | 0.732 | 1.161 | 1.457 | 2.093 |
Kit components
| Catalog number | Description | Quantity | Storage of opened/reconstituted material |
|---|---|---|---|
| EZ1765-CA | Mouse anti-human PAM monoclonal antibody (Capture Antibody) | 500 µl, 4 µg/mL (recommended dilution 1:100) | Store undiluted at 4°C for 1 month or at -20°C for 3 months provided this is within the expiration date of the kit. |
| EZ1765-DA | Biotinylated goat anti-human PAM polyclonal antibody (Detection Antibody) | 500 µl (recommended dilution 1:100) | |
| AR1103 | Avidin-Biotin-Peroxidase Complex (ABC) | 500 µl (recommended dilution 1:100) | |
| EZ1765-ST | Lyophilized recombinant human PAM standard | 10 ng/tube×3 | Discard the standard stock solution after 12 hours at 4°C. May be stored at -20°C for 48 hours provided this is within the expiration date of the kit. |
Materials required but not provided
- Microplate reader in standard size.
- Automated plate washer.
- Incubator.
- Adjustable pipettes and pipette tips. Multichannel pipettes are recommended in the condition of large amount of samples in the detection.
- Clean tubes and Eppendorf tubes.
- 96 well microplate (Cat# AR1100)
- Plate Sealers.
- Capture Antibody Diluent: PBS.
- Reagent Diluent: 1% BSA in PBS, pH 7.2-7.4, 0.2 µm filtered.
- Substrate Solution: Tetramethylbenzidine (Cat# AR1104)
- Stop Solution: 2 N H2SO4 (Cat# AR1105)
- Wash Buffer (PBS and PBS-T). Recipes:
- PBS: 8g NaCl, 0.2g KCl, 1.15g Na2HPO4, 0.2g KH2PO4, adjust the total volume to 1 L with distilled water, pH 7.2-7.4, 0.2 µm filtered.
- PBS-T: 0.1% Tween® 20 in PBS, pH 7.2-7.4.
- *Item 6 – 12 are included in the EZ Set Accessory Kit (EZA001)
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