Alkaline Phosphatase antibody | AbD25296_hIgG2
A microtiter plate was coated overnight with various antigens at a concentration of 5 μg/ml. After washing and blocking with PBST+5% BSA, detection was performed using Human Anti-Bacterial Alkaline Phosphatase:HRP Antibody (clone AbD25296_hIgG2) at a concentration of 2 μg/ml followed by Mouse Anti-Human IgG (Fc) CH2 Domain:HRP Antibody (MCA647P) in HISPEC Assay Diluent (BUF049A) and QuantaBlu Fluorogenic Peroxidase Substrate.
Western blot analysis was performed by loading 25 μl Hela lysate (lanes 2 and 3) and Precision Plus Protein™ Prestained Standards (lane 1) onto a AnykD™ Criterion™ TGX Stain-Free™ Protein Gel. The gel was run under reducing conditions and proteins were transferred to a nitrocellulose membrane. Lane 2 was probed with Anti-HSPA8 Antibody, clone AbD28028 Fab-A-FH format, at a concentration of 10 μg/ml. Lanes 2 and 3 were then probed with Human Anti-Bacterial Alkaline Phosphatase:HRP (clone AbD25296_hIgG2, HCA275P) at dilution of 1/1000. Visualization was carried out using the ChemiDoc™ MP Imaging System with an auto exposure. The Western blot bands (red) and protein standards (green) have been pseudocolored. Fab-A-FH format is a bivalent bacterial alkaline phosphatase fusion antibody (DYKDDDDK- and His-6-tags).
A microtiter plate was coated overnight with recombinant GFP antibody (His-tagged) at a concentration of 5 μg/ml. After washing and blocking with PBST+5% BSA, detection was performed using primary human anti-GFP antibodies in different bivalent Fab formats (e.g. Fab-A-V5Sx2) titrated to the given concentrations in PBST, followed by Human Anti-Bacterial Alkaline Phosphatase:HRP Antibody (clone AbD25296_hIgG2) at a concentration of 2 μg/ml in HISPEC Assay Diluent (BUF049A) and QuantaBlu Fluorogenic Peroxidase Substrate. Data are shown as the mean of triplicate measurements.
Figure A (control experiment). Mouse Anti-Human CD4:Alexa Fluor 647 (MCA1267A647) and Mouse Anti-Human CD3 (MCA2184) detected by Human Anti-Mouse IgG2a, (clone AbD24124, bivalent Fab format bacterial alkaline phosphatase (BAP) fusion antibody), followed by FITC conjugated isotype control, Recombinant Human IgG2 Lambda (HCA108). FITC conjugation of the isotype control antibody was performed using a LYNX Rapid Fluorescein Antibody Conjugation Kit (LNK063F). Figure B. Mouse Anti-Human CD4:Alexa Fluor 647 (MCA1267A647) and Mouse Anti-Human CD3 (MCA2184) detected by Anti-Mouse IgG2a (clone AbD24124, bivalent format Fab bacterial alkaline phosphatase (BAP) fusion antibody) followed by Human Anti-Bacterial Alkaline Phosphatase:FITC (HCA275F). All experiments performed on red blood cell lysed human blood, gated on live, single cell lymphocytes in the presence of 10% human serum. Data acquired on the ZE5 Cell Analyzer.
Human anti Bacterial Alkaline Phosphatase:FITC
This FITC conjugated Anti-Bacterial Alkaline Phosphatase Antibody is recommended for use as a secondary detection reagent with HuCAL antibodies in Fab-A and Fab-Max formats.
Human anti Bacterial Alkaline Phosphatase:HRP
This HRP conjugated Anti-Bacterial Alkaline Phosphatase Antibody is recommended for use as a secondary detection reagent with HuCAL antibodies in Fab-A and Fab-Max formats.
- Product Type
- Monoclonal Antibody
- Clone
- AbD25296_hIgG2
- Isotype
- IgG2
| Product Code | Applications | Pack Size | List Price | Quantity |
|---|---|---|---|---|
| HCA275F | F | 0.1 mg |
|
|
| HCA275P | E WB | 0.1 mg |
|
Human Anti-Bacterial Alkaline Phosphatase Antibody, clone AbD25296_hIgG2 recognizes bacterial alkaline phosphatase (BAP), a periplasmic enzyme which catalyzes the release of 5'- and 3'-phosphate groups from DNA, RNA and nucleotides. BAP can also dephosphorylate a number of proteins and alkaloids, and has an optimum pH of 8.0. BAP is encoded by the phoA gene, and forms a homodimer of two identical ~47 kDa subunits.
Fab-A and Fab-Max format antibodies generated using HuCAL® technology are formed via dimerization of BAP. The Anti-BAP Antibody HCA275P can be used as a secondary detection reagent with HuCAL antibodies in Fab-A and Fab-Max formats (Fab-Max is a modified BAP fusion antibody with inactivated enzymatic activity).
View further information about HuCAL antibody formats.
Fab-A and Fab-Max format antibodies generated using HuCAL® technology are formed via dimerization of BAP. The Anti-BAP Antibody HCA275P can be used as a secondary detection reagent with HuCAL antibodies in Fab-A and Fab-Max formats (Fab-Max is a modified BAP fusion antibody with inactivated enzymatic activity).
View further information about HuCAL antibody formats.
Product Details
- Target Species
- Bacterial
- Product Form
- Human IgG2 antibody (lambda light chain) selected from the HuCAL® phage display library and expressed in a human cell line. Conjugated to Fluorescein Isothiocyanate Isomer 1 (FITC) - liquid
- Product Form
- Human IgG2 antibody (lambda light chain) selected from the HuCAL® phage display library and expressed in a human cell line. Conjugated to horseradish peroxidase (HRP) - liquid
- Preparation
- Purified IgG prepared by affinity chromatography on Protein A
- Preparation
- Purified IgG prepared by affinity chromatography on Protein A
- Buffer Solution
- Phosphate buffered saline
- Buffer Solution
- Phosphate buffered saline
- Preservative Stabilisers
- 0.09% Sodium Azide (NaN3)
1% Bovine Serum Albumin - Preservative Stabilisers
- 0.01% Thiomersal
HRP Stabiliser (BUF052A) - Immunogen
- Bacterial alkaline phosphatase
- Affinity
- The monovalent intrinsic affinity of this antibody was measured as KD=38pM by real time, label-free molecular interaction analysis on immobilized HuCAL antibody in the Fab-A-FH format
- Approx. Protein Concentrations
- IgG concentration 0.1 mg/ml
- Approx. Protein Concentrations
- IgG concentration 1.0 mg/ml
Storage Information
- Storage
- This product is shipped at ambient temperature. It is recommended to aliquot and store at -20°C on receipt. When thawed, aliquot the sample as needed. Keep aliquots at 2-8°C for short term use (up to 4 weeks) and store the remaining aliquots at -20°C.
Avoid repeated freezing and thawing as this may denature the antibody. Storage in frost-free freezers is not recommended. - Storage
- Store at +4oC. DO NOT FREEZE
This product should be stored undiluted. This product is photosensitive and should be protected from light. Should this product contain a precipitate we recommend microcentrifugation before use - Guarantee
- 12 months from date of despatch
- Guarantee
- 12 months from date of despatch
More Information
- UniProt
- P00634
- GO Terms
- GO:0005515 protein binding
- GO:0000287 magnesium ion binding
- GO:0004035 alkaline phosphatase activity
- GO:0008270 zinc ion binding
- GO:0030613 oxidoreductase activity, acting on phosphorus or arsenic in donors
- GO:0033748 hydrogenase (acceptor) activity
- GO:0042597 periplasmic space
- Acknowledgements
- Sold under license of U.S. Patents 7785859 and 8273688 and corresponding patents. This antibody was developed by Bio-Rad.
- Acknowledgements
- Sold under license of U.S. Patents 7785859 and 8273688 and corresponding patents. This antibody was developed by Bio-Rad.
- Regulatory
- For research purposes only
Applications of Alkaline Phosphatase antibody
| Application Name | Verified | Min Dilution | Max Dilution |
|---|---|---|---|
| Flow Cytometry | 1/5 | 1/50 | |
| ELISA | |||
| Western Blotting | 1/1,000 | 1/10,000 |
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
- Technical Advice
- Recommended protocols and further information about HuCAL recombinant antibody technology can be found in the HuCAL Antibodies Technical Manual
- Technical Advice
- Recommended protocols and further information about HuCAL recombinant antibody technology can be found in the HuCAL Antibodies Technical Manual
Copyright © 2021 Bio-Rad Antibodies (formerly AbD Serotec)
Negative Isotype Controls Available
| Description | Product Code | Applications | Pack Size | List Price | Quantity |
|---|---|---|---|---|---|
| Recombinant Human IgG2 Lambda | HCA108 | E | 0.1 mg |
|
Useful Reagents Available
| Description | Product Code | Applications | Pack Size | List Price | Quantity |
|---|---|---|---|---|---|
| Hispec Assay Diluent | BUF049A | E IY | 50 ml | ||
| LYNX Rapid Fluorescein Antibody Conjugation Kit | LNK063F | CJ | 3 Conjugations For 20µg Antibody | ||
| Mouse anti Human CD4:Alexa Fluor® 647 | MCA1267A647 | F | 100 Tests/1ml |
|
|
| Mouse anti Human CD3 | MCA2184 | F IP | 0.2 mg |
|
|
| Hispec Assay Diluent | BUF049A | E IY | 50 ml |
Product Specific References
References for Alkaline Phosphatase antibody
-
Harth, S. et al. (2019) Generation by phage display and characterization of drug-target complex-specific antibodies for pharmacokinetic analysis of biotherapeutics.
MAbs. 11 (1): 178-90.
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