Complement Factor Bb 抗体
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Quick Overview for Complement Factor Bb 抗体 (ABIN2473066)
抗原
适用
宿主
克隆类型
标记
应用范围
克隆位点
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特异性
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Specificity property: neoantigen
The monoclonal antibody, clone 032B-22.1.X, binds to a neoantigen expressed on the Bb fragment of human Factor B. This antigen is not expressed on native Factor B and, accordingly, the antibody does not bind to native Factor B found in human plasma. -
产品特性
- Purified IgG
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纯化方法
- Purified
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亚型
- IgG2a
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anti-Complement Factor Bb antibody (APC)
适用: 人 FACS, ICC, IF, IHC 宿主: 兔 Polyclonal APC
anti-Complement Factor Bb antibody (PE)适用: 人 FACS, ICC, IF, IHC 宿主: 兔 Polyclonal PE
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应用备注
- Optimal working dilution should be determined by the investigator.
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说明
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If the Factor B is denatured in any way, the protein can express neoantigen(s) that are not present in functionally active, native Factor B. For example, if the Factor B is bound to a well in a microtiter plate, a portion of it will be conformationally distorted and express the Bb neoantigen. Purification of Factor B can also result in denaturation of a portion of the Factor B and this denatured protein can also express the Bb-specific neoantigen. Finally, lack of careful handling of serum and plasma can result in complement activation and generation of small amounts of Bb and Ba fragments; in this case low levels of Bb will be detectable with this antibody and with the MicroVue assay for Bb.
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限制
- 仅限研究用
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- by
- Complement Physiopathology, Departamento de Microbiología I, Universidad Complutense de Madrid
- No.
- #103567
- 日期
- 2019.02.22
- 抗原
- Bb
- Lot Number
- 146096
- Method validated
- Western Blotting
- Positive Control
Purified Bb fragment and zymosan activated serum
- Negative Control
Non-activated sera
- Notes
ABIN2473066 recognizes both Factor B and Bb fragment in plasma samples. It does not specifically recognize a neo-epitope in Bb fragment.
- Primary Antibody
- ABIN2473066
- Secondary Antibody
- goat anti-mouse Immunoglobulins/HRP (Dako, P0447, lot 20058518)
- Full Protocol
- Purify Bb fragment from plasma samples.
- Separate 10ng purified Bb fragment and 0.5µl of zymosan activated serum and 0,5 µl of control serums on a 10% SDS-PAGE electrophoresis gel for 1h 120 V.
- Activate PVDF membrane with methanol for 15sec and rinse with transfer buffer for 5min. Transfer proteins from the gel to the PVDF membrane using a Mini Trans-Blot cell Tetra Blotting Module (Bio-Rad, 1660827EDU) for 1h at 100V.
- Stain membrane using Ponceau S to verify protein transfer.
- Block the membrane with blocking buffer (1x PBS, 3% BSA, 0.1% Tween) for 30min at RT.
- Incubate membrane with primary mouse anti-complement factor Bb antibody (antibodies-online, ABIN5692886, lot 146096) diluted 1:1000 in working buffer (1x PBS 1% BSA, 0.1% Tween) ON at 4°C.
- Wash membrane 3x for 5min with PBST (1x PBS, 0.1% Tween).
- Incubate membrane with secondary goat anti-mouse Immunoglobulins/HRP (Dako, P0447, lot 20058518) diluted 1:1000 in working buffer for 30min at RT.
- Wash membrane 3x for 5min with PBST.
- Reveal protein bands using ClarityTM Western ECL substrate (Bio-Rad, 170-5061, lot 102031078) on a ChemiDoc MP Imaging System (Bio-Rad, 17001402).
- Experimental Notes
In dentatured samples the complement Factor Bb antibody directed against a neo-epitope in Bb fragment ABIN2473066 reveals two proteins of the expected molecular weight of Factor B (90kDa) and Bb fragment (60kDa). The antibody does not specifically recognize a neo-epitope in Bb fragment and is thus not suitable for selectively detecting the Bb fragment subsequently to denaturation.
As an additional control we performed a Western Blot using in-house antibodies which detect Factor B and fragments Bb and Ba (not shown).
生效 #103567 (Western Blotting)!['独立验证'标志]( "成功验证")
!['独立验证'标志]( "成功验证")
Validation Images![Western blot analysis of purified Bb fragment (1), zymosan activated serum (2), and non-activated sera (3, 4, 5) using ABIN5692886.]()
Western blot analysis of purified Bb fragment (1), zymosan activated serum (2), and non-activated sera (3, 4, 5) using ABIN5692886.
Full Methods -
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状态
- Liquid
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浓度
- 1.0 mg/mL
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: "Structural biology of the alternative pathway convertase." in: Immunological reviews, Vol. 180, pp. 123-35, (2001) (PubMed).
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: "Structural biology of the alternative pathway convertase." in: Immunological reviews, Vol. 180, pp. 123-35, (2001) (PubMed).
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- Complement Factor Bb
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基因ID
- 629
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UniProt
- P00751
抗原
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