Recombinant Mouse Anti-AAV VP1 Antibody scFv Fragment (A20) (CAT#: PSBL-390)
Recombinant Mouse Antibody (A20) scFv Fragment is specific to AAV VP1, expressed in E. coli. This scFv fragment neutralizes AAV-2 and AAV-3B subsequent to primary receptor binding.
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Figure 1 Influence of mutation of pore-forming amino acids on genome packaging.
(B) Capsids were immunoprecipitated (IP) from frozen-thawed lysates obtained from 293T cells transfected with a plasmid harboring an AAV2 wt or mutated genome and superinfected with Ad5 (MOI, 2) by protein A-Sepharose-bound antibody A20. Control (C) precipitations were also performed with a non-AAV-related monoclonal antibody. Samples were digested (+) or not digested (−) with DNase I to discriminate between capsid-associated and packaged DNAs. Coprecipitated genomes were isolated and electrophoresed on neutral agarose gels. Southern blotting was performed, and genomes were detected with a rep-specific probe. A 4.7-kb fragment was used as a marker (top panels, M). A portion of each immunoprecipitate was tested by Western blot analysis for the level of capsid recovery; an extract from Ad5- and AAV2-coinfected HeLa cells was used as a marker (bottom panels, M).
Bleker, S., Sonntag, F., & Kleinschmidt, J. A. (2005). Mutational analysis of narrow pores at the fivefold symmetry axes of adeno-associated virus type 2 capsids reveals a dual role in genome packaging and activation of phospholipase A2 activity. Journal of virology, 79(4), 2528-2540.
Figure 2 Externalization of VP1/VP2 N termini during infection.
(B) Analysis of cell lysates. HeLa cells were incubated with wt AAV2 (10,000 particles/cell) for 30 min at 4°C and washed. Following incubation at 37°C, the cells were harvested and lysed by freezing-thawing at the indicated time points. Virus-containing supernatant was analyzed by a native immuno-dot blot using VP1-specific antibody A1, VP1/VP2, specific antibody A69, C-terminus-specific antibody B1, or capsid-specific antibody A20. Extracts of noninfected cells were used as a negative control (n). Signal intensities were measured with Image Quant TL software, and the ratios of A1/A20 and A69/A20 signal were determined to quantify exposure of N termini on capsids. Means ± standard deviations from three independent experiments are shown.
Sonntag, F., Bleker, S., Leuchs, B., Fischer, R., & Kleinschmidt, J. A. (2006). Adeno-associated virus type 2 capsids with externalized VP1/VP2 trafficking domains are generated prior to passage through the cytoplasm and are maintained until uncoating occurs in the nucleus. Journal of virology, 80(22), 11040-11054.
Figure 3 AAV2 enters the cytoplasm with exposed VP1/VP2 N termini.
(A) Extracellular neutralization of infection. AAV2 particles were preincubated with 100 μg/ml of VP1-specific antibody A1, VP1/VP2-specific antibody A69, C-terminus-specific antibody B1, or capsid-specific antibody A20. In a control experiment, no antibody (no Ab) was added. The samples were transferred to HeLa cells (2 × 10³ particles/cell), infected with Ad5 (MOI = 4), and incubated for 20 h. Early viral gene expression was analyzed by immunofluorescence with Rep-specific antibody 76.3. Cell nuclei were visualized with DAPI. Means ± standard deviations from two independent experiments are shown.
Sonntag, F., Bleker, S., Leuchs, B., Fischer, R., & Kleinschmidt, J. A. (2006). Adeno-associated virus type 2 capsids with externalized VP1/VP2 trafficking domains are generated prior to passage through the cytoplasm and are maintained until uncoating occurs in the nucleus. Journal of virology, 80(22), 11040-11054.
Specifications
- Immunogen
- AAV Capsid protein VP1
- Host Species
- Mouse
- Derivation
- Mouse
- Type
- scFv
- Specificity
- Tested positive against native AAV VP1
- Species Reactivity
- AAV
- Clone
- A20
- Applications
- WB, ELISA, FuncS, IP, DB, Neut
Product Property
- Purity
- >95% by SDS-PAGE and HPLC analysis
- Storage
- Store the antibody (in aliquots) at -20°C. Avoid repeated freezing and thawing of samples.
Applications
- Application Notes
- The VP1 antibody has been reported in applications of Western Blot, Enzyme-linked Immunosorbent Assay, Functional Assay, Immunoprecipitation, Dot Blot, Neutralization.
For Neutralization: wt AAV2 particles (2.5 × 10⁸) were incubated with 100 μg/ml purified antibody A1, A69, B1, or A20 in a total volume of 5 μl (PBS) for 30 min at room temperature. Then, the samples were diluted in 1 ml medium and added in combination with Ad5 (MOI = 4) to 1.4 × 10⁵ HeLa cells grown overnight on coverslides. Rep expression was analyzed after 20 h as described above.
Target
Product Notes
This is a product of Creative Biolabs' Hi-Affi™ recombinant antibody portfolio, which has several benefits including:
• Increased sensitivity
• Confirmed specificity
• High repeatability
• Excellent batch-to-batch consistency
• Sustainable supply
• Animal-free production
See more details about Hi-Affi™ recombinant antibody benefits.
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PABL-090 | Recombinant Mouse Anti-EV71 VP1 Antibody (MA28-7) | WB, ELISA, Neut, FuncS | IgG |
PABZ-047 | Recombinant Mouse Anti-FMDV VP1 pentadecapeptide (YTTSTRGDLAHVTAT) Antibody (4C4) | Neut, ELISA | IgG |
PABW-047 | Recombinant Mouse Anti-FMDV-C-S8c1 VP1 Antibody (SD6) | Neut | IgG |
FAMAB-0138-CN | Recombinant Human Anti-VP1 Antibody (1418-1) | WB, ELISA, FC, IP | Human IgG1, Κ |
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No Non-Specific Binding
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For Research Use Only. Not For Clinical Use.
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