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.
Filter by Application:
E WB ResetHuman anti Bacterial Alkaline Phosphatase:FITC
- Product Type
- Monoclonal Antibody
- Clone
- AbD25296_hIgG2
- Isotype
- IgG2
- Specificity
- Alkaline Phosphatase
Antibody Quality and 
Unrivaled 
| 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. |
- 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
- Preparation
- Purified IgG prepared by affinity chromatography on Protein A
- Buffer Solution
- Phosphate buffered saline
- Preservative Stabilisers
- 0.09% Sodium Azide (NaN3)
1% Bovine Serum Albumin - 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
- Max Ex/Em
-
Fluorophore Excitation Max (nm) Emission Max (nm) FITC 490 525 - Regulatory
- For research purposes only
- Guarantee
- 12 months from date of despatch
- Acknowledgements
- This product and/or its use is covered by claims of U.S. patents, and/or pending U.S. and non-U.S. patent applications owned by or under license to Bio-Rad Laboratories, Inc. See bio-rad.com/en-us/trademarks for details.
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.
Avoid repeated freezing and thawing as this may denature the antibody. Storage in frost-free freezers is not recommended.
This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visit the antibody protocols page.
| Application Name | Verified | Min Dilution | Max Dilution |
|---|---|---|---|
| Flow Cytometry |  |
1/5 | 1/50 |
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
| Description | Product Code | Applications | Pack Size | List Price | Your Price | Quantity | |
|---|---|---|---|---|---|---|---|
| Recombinant Human IgG2 Lambda | HCA108 | E | 0.1 mg |
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| List Price | Your Price | ||||||
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| Description | Recombinant Human IgG2 Lambda | ||||||
| Description | Product Code | Applications | Pack Size | List Price | Your Price | Quantity | |
|---|---|---|---|---|---|---|---|
| Hispec Assay Diluent | BUF049A | E IY | 50 ml | Log in | |||
| List Price | Your Price | ||||||
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| Description | Hispec Assay Diluent | ||||||
| LYNX Rapid Fluorescein Antibody Conjugation Kit | LNK063F | CJ | 3 Conjugations For 20µg Antibody | Log in | |||
| List Price | Your Price | ||||||
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| Description | LYNX Rapid Fluorescein Antibody Conjugation Kit | ||||||
| Mouse anti Human CD4:Alexa Fluor® 647 | MCA1267A647 | F | 100 Tests/1ml |
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| List Price | Your Price | ||||||
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| Description | Mouse anti Human CD4:Alexa Fluor® 647 | ||||||
| Mouse anti Human CD3 | MCA2184 | F IP | 0.2 mg |
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| List Price | Your Price | ||||||
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| Description | Mouse anti Human CD3 | ||||||
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.
- 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
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