Immunoprecipitation Kit (Protein A Agarose Gel)
mRNA synthesis
In vitro transcription of capped mRNA with modified nucleotides and Poly(A) tail
Tyramide Signal Amplification (TSA)
TSA (Tyramide Signal Amplification), used for signal amplification of ISH, IHC and IC etc.
Phos Binding Reagent Acrylamide
Separation of phosphorylated and non-phosphorylated proteins without phospho-specific antibody
Cell Counting Kit-8 (CCK-8)
A convenient and sensitive way for cell proliferation assay and cytotoxicity assay
SYBR Safe DNA Gel Stain
Safe and sensitive stain for visualization of DNA or RNA in agarose or acrylamide gels.
Inhibitor Cocktails
Protect the integrity of proteins from multiple proteases and phosphatases for different applications.
免疫沉淀(Immunoprecipitation,IP)或免疫共沉淀(Co-Immunoprecipitation,Co-IP)是研究蛋白或蛋白与蛋白间相互作用(Protein-Protein Interactions, PPIs)的常用技术手段,通过使用特异性抗体和可结合抗体的介质(如Protein A/G琼脂糖凝胶等),或直接使用偶联特异性抗体的介质(如琼脂糖凝胶或磁珠),将目标蛋白质从复杂样品中分离出来,后续可用于Western印迹检测或质谱分析等。
Protein A是一种发现于金黄色葡萄球菌(Staphylococcus aureus)的细胞壁表面蛋白,分子量为42 kDa,能特异性地与哺乳动物免疫球蛋白(Immunoglobulin, Ig) Fc区结合,同时也会和人VH3家族的Fab区结合。Protein A可特异性地结合相应抗体,如human IgG1、IgG2、IgG4,mouse IgG2a、IgG2b及rabbit IgG等,且每个Protein A分子可以结合5个IgG分子。针对常见的免疫球蛋白亚类的结合能力及不同物种的总结合能力情况见下表(Table 1)。
本试剂盒包含Protein A琼脂糖凝胶及其他免疫沉淀必要试剂,配合用户自备的特异性抗体,使免疫沉淀(Immunoprecipitation, IP,也称Pull-down)或免疫共沉淀(Co-IP)实验更加简单、便捷、高效,免疫沉淀产物可用于目的蛋白或其蛋白复合物的检测等实验。(具体产品参数见Table 2)
Species |
Subclass |
Protein A binding |
Protein G binding |
Human |
lgA |
++ |
- |
lgD |
++ |
- |
|
IgE |
++ |
- |
|
lgG1 |
++++ |
++++ |
|
lgG2 |
++++ |
++++ |
|
IgG3 |
- |
++++ |
|
lgG4 |
++++ |
++++ |
|
lgM |
++ |
- |
|
Mouse |
IgG1 |
+ |
++++ |
lgG2a |
++++ |
++++ |
|
lgG2b |
+++ |
+++ |
|
lgG3 |
++ |
+++ |
|
lgM |
+/- |
- |
|
Rat |
lgG1 |
- |
+ |
lgG2a |
- |
++++ |
|
lgG2b |
- |
++ |
|
lgG3 |
+ |
++ |
|
Avian egg yolk |
IgY |
- |
- |
Cow |
|
++ |
++++ |
Dog |
|
++ |
+ |
Goat |
|
- |
++ |
Guinea Pig |
lgG1 |
++++ |
++ |
lgG2 |
++++ |
++ |
|
Hamster |
|
+ |
++ |
Horse |
|
++ |
++++ |
Koala |
|
- |
+ |
Llama |
|
- |
+ |
Monkey (rhesus) |
|
++++ |
++++ |
Pig |
|
+++ |
+++ |
Rabbit |
|
++++ |
+++ |
Sheep |
|
+/- |
++ |
Table 1 Affinity data for Protein A for different sources and subtypes of IgG. ++++ = Strong Binding; ++~+++ = Medium Binding; + = Weak Binding; +/- = Weak or No Binding; - = No Binding.
Characteristics |
Description |
Product content |
Store in 20% ethanol with a glue-to-suspension ratio of approximately 75% |
Bead structure |
4% cross-linked agarose |
Average beads size |
90 μm |
Coupled protein |
Recombinant Protein A (E.coli) |
M. W. of protein |
14 kDa Protein A |
Ligand concentration |
6 mg rProtein A per mL settled gel |
Binding capacity |
~35 mg human IgG per ml settled gel |
Elution method |
Elution with acid, competing peptide or SDS-PAGE loading buffer |
Operating temperature |
2-40℃ |
pH stability* |
3-10 |
Solvent resistant |
6 M guanidine hydrochloride, 8 M urea, 2% benzyl alcohol**, 75% ethanol**, 1 mM NaOH (pH 11)., 0.1 M sodium citrate/HCl (pH 3). |
Application |
Suitable for IP, Co-IP, Protein purification |
Storage |
4°C |
*After 7 days of storage at 40°C, pH3-10, the physicochemical properties, and functions of agarose showed no significant changes. ** v/v, volume ratio |
Table 2 The main related indicators of Protein A Agarose Gel.
Components | K4621-20 T |
Cell Lysis Buffer | 50 mL |
Protease Inhibitor Cocktail (EDTA-Free,100X in DMSO) | 0.5 mL |
10X TBS | 10 mL |
Neutralization Buffer | 200 μL |
Acid Elution Buffer | 2 mL |
Protein A Agarose Gel | 400 μL |
5X Protein Loading Buffer (Reducing) | 600 μL |
Store Protease Inhibitor Cocktail (EDTA-Free,100X in DMSO) and 5X Protein Loading Buffer (Reducing) at -20°C and the rest of components at 4°C for 12 months. |