产品名称：RAB-IP Anti-rabbit IgG (HRP) for IP/CoIP

货号： IF9203

品牌：Engibody

简介：IP/CoIP专用二抗，消除抗体分子轻重链干扰

RAB-IP™ IP专用的抗兔IgG二抗（天然构象完整分子抗体特异性）（HRP偶联）是一种蛋白IP-WB试剂，其能够无障碍地检测来自上游IP/CoIP的蛋白质印迹目标蛋白带，而不受变性IgG的干扰（IP一抗）。这允许在不受IgG重链（50kDa）和轻链（25kDa）干扰的情况下检测（共）免疫沉淀蛋白。通常，这种干扰倾向于源于传统的二抗，其识别在免疫沉淀过程中与抗原一起释放的一抗或来自裂解物本身的内源性IgG的轻链和重链。

RAB-IP™ 如果免疫沉淀的抗原-抗体复合物在下游WB之前被还原和变性，则用于IP的抗兔IgG（HRP标记）仅识别天然（非还原）一抗（来自兔），因此重链和轻链的条带不会显示。

描述

RAB-IP™ IP专用的抗兔IgG二抗（天然构象完整分子抗体特异性）（HRP偶联）

反应性

识别兔IgG完整抗体分子

经过测试的应用实验

WB:1/1000-1/2000。最佳稀释度应由最终用户确定。

特异性

该二抗对天然兔rabbit多克隆/单克隆抗体完整分子具有特异性，同时不识别兔IgG抗体单独轻重链

免疫原

以天然兔IgG为免疫原，在山羊身上制备抗体，然后制备只识别兔抗体完整分子同时不识别兔抗体单独轻重链的单克隆抗体。

克隆性

单克隆，克隆号：3B8

同型对照

山羊IgG

类型

抗体在10 mM PBS中，pH 7.4，50%甘油，10 mg/mL（1%w/v）BSA作为稳定剂，0.01%（w/v）硫柳汞作为防腐剂。

存储说明

储存于-20°C，避免冷冻/解冻循环。不要分装抗体。

偶联标记

辣根过氧化物酶（HRP）

抗体来源宿主

山羊

RAB-IP™ Anti- rabbit IgG for IP (HRP Conjugated) only recognize native (non-reduced) primary antibodies (from rabbit) and therefore the bands of heavy and light chains is highly minimized, if the immunoprecipitated antigen-antibody complex is fully reduced and denatured before downstream WB.

• Description

• RAB-IP™ Anti- rabbit IgG (native conformation antibody specific) for IP (HRP Conjugated)

• Reactivity

• Rabbit

• Tested applications

• WB : 1/1000-1/2000. Optimal dilutions should be determined by the end user.

• Specificity

• This secondary antibody is specific to native rabbit polyclonal/monoclonal antibody

• Immunogen

• The antibody was developed in goat using the nature rabbit IgG as the immunogen and then develop monoclonal antibody.

• Clonality

• Monoclonal, clone number: 3B8

• Isotype

• Goat IgG

• Form

• The antibody in 10 mM phosphate buffered saline (PBS), pH 7.4, 50% glycerol with 10 mg/mL (1% w/v) BSA (IgG free, protease free) as a stabilizer and 0.01% (w/v) thimerosal as preservative.

• Storage instruction

• Store at -20°C, Avoid freeze / thaw cycle. Do not aliquot the antibody.

• Conjugation

• Horseradish Peroxidase (HRP)

• Host

• Goat

Figure 1. RAB-IP™ Anti- rabbit IgG for IP (HRP) secondary antibody images: Western blotting.

IP conditions:

target protein TBP was immunoprecipitated from 0.5 mL cell lysate of 1x107 Hela cells with 5 µg anti-human TBP rabbit monoclonal antibody and protein A/G agarose beads (Engibody, IF0001).

WB conditions:

the immunoprecipitated antigen-antibody complex should be boiled for 5-10 minutes in SDS sample buffer with a increase in SDS amount if required, make sure that the complex is fully reduced and denatured before downstream WB.

then electrophoresis, transferred to a PVDF membrane, and incubate with an anti- TBP rabbit monoclonal antibody.

Secondary Antibody Detection:

Lane 1: Detection with RAB-IP™ Anti-rabbit IgG for IP (HRP) (CAT: IF9203)

The heavy and light chains can not be seen, confirming that although the immunoprecipitating heavy and light-chains are present, detects only native antibody and not denatured heavy and light-chains.

Lane 2: Detection with a traditional goat anti-rabbit IgG (H&L) (HRP) secondary antibody (CAT: AT0097)

Figure 2. A schematic representation of IP-WB workflow by using RAB-IP™ secondary antibody.

Using primary antibody coupled beads

RAB-IP™ Anti-rabbit IgG for IP (HRP Conjugated) only recognize native (non-reduced) primary antibodies (from rabbit) and therefore the bands of heavy chain and light chain are highly minimized, if the immunoprecipitated antigen-antibody complex is fully reduced and denatured before downstream WB.

Figure 3. A schematic representation of IP-WB workflow by using RAB-IP™ secondary antibody.

Using protein A/G Beads + primary antibody

RAB-IP™ Anti-rabbit IgG for IP (HRP Conjugated) only recognize native (non-reduced) primary antibodies (from rabbit) and therefore the bands of heavy chain and light chain are highly minimized, if the immunoprecipitated antigen-antibody complex is fully reduced and denatured before downstream WB.