Universal Magnetic Co-IP Kit Overview
While Co-IP is often used to study cytoplasmic protein complexes, traditional methods are not optimal for studying DNA-binding proteins because these nuclear complexes are fragile and can be difficult to remove from the DNA. This causes them to either be disrupted during extraction, or to be lost because they remain bound to the DNA. For this reason, in addition to containing components for whole-cell extraction, the Universal Magnetic Co-IP Kit provides nuclear extraction reagents that have been optimized to preserve nuclear protein complexes.
Universal Magnetic Co-IP Kit Contents
The Universal Magnetic Co-IP Kit is shipped on dry ice and contains reagents with multiple storage temperatures inside. Please store components according to the storage conditions in the manual after first use. All reagents are guaranteed stable for 6 months from date of receipt when stored properly.
- 10X PBS, 4°C
- 10X Hypotonic Buffer, 4°C
- Whole-cell Lysis Buffer, -20°C
- Co-IP/Wash Buffer, 4°C
- Detergent, 4°C
- 100X Phosphatase Inhibitors, -20°C
- Protease Inhibitor Cocktail, -20°C
- Deacetylase Inhibitor, -20°C
- 100 mM PMSF, -20°C
- 0.5 M EDTA, 4°C
- Digestion Buffer, 4°C
- Enzymatic Shearing Cocktail, -20°C
- Protein G Magnetic Beads, 4°C
- Bar Magnet, RT
- Mini Glue Dots, RT
Universal Magnetic Co-IP Kit Data
Universal Magnetic Co-IP Kit FAQs
What is the difference between the Nuclear Complex Co-IP Kit (Cat No. 54001) and the Universal Magnetic Co-IP Kit (Cat No. 54002)?
Below are the main differences:
|Nuclear Complex Co-IP Kit (Cat No. 54001)||Universal Magnetic Co-IP kit (Cat No. 54002)|
|Processes nuclear lysates||Processes nuclear and whole-cell lysates|
|More customizable offering varying stringencies||Easier to use|
|Uses Protein G agarose columns
||Uses magnetic beads (included in kit)|
Should I increase the IP stringency when using the Universal Magnetic Co-IP Kit?
The Protein G Magnetic Beads included in the Universal Magnetic Co-IP Kit were chosen because they exhibit a very small degree of non-specific binding. This helps ensure very low background, enabling the Complete Co-IP/Wash Buffer used in the kit to be of low stringency, which better preserves fragile protein complexes. Particular cell lines, stimulations or antibodies may result in higher background. If your experiment has more background than is optimal, we recommend increasing the number of washes performed from 4 to 6.
Is this kit compatible with downstream mass spectrometry (MS/LC-MS)?
Active Motif has not used this kit with downstream mass spectrometry. However, there are many publications where it has been used for this application. Various MS instruments are sensitive to different buffer components. Please consult with your instrument’s manual or core facility to determine any components to avoid and we can let you know if they are present in our buffer systems.
Universal Magnetic Co-IP Kit Publications
Search our database of customer publications that have used our Universal Magnetic Co-IP Kit.
Universal Magnetic Co-IP Kit Documents
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|Universal Magnetic Co-IP Kit||25 rxns||54002||$460||Buy|
Universal Magnetic Co-IP Kit advantages
- Perform Co-IP of nuclear or cytoplasmic protein complexes, beginning from cells or tissue
- Magnetic beads speed and simplify the procedure while reducing background, enabling the use of low-stringency buffers that maintain complex integrity
- Nuclear extraction procedure removes nuclear protein complexes from the DNA without disrupting protein/protein interactions
- Inhibitor solutions included to preserve the integrity of protein modifications
Perform Co-IP of cytoplasmic & nuclear protein complexes
While Co-IP is often used to study cytoplasmic protein complexes, traditional methods are not optimal for studying DNA-binding proteins because these nuclear complexes are fragile and can be difficult to remove from the DNA. This causes them to either be disrupted during extraction, or to be lost because they remain bound to the DNA. For this reason, in addition to containing components for whole-cell extraction, the Universal Magnetic Co-IP Kit provides nuclear extraction reagents that have been optimized to preserve nuclear protein complexes. The reduced background provided by the low non-specific binding of the kit's magnetic beads makes it possible to use a low-stringency Co-IP/Wash Buffer that helps maintain the protein complexes (Figure 1). To further improve Co-IP of nuclear complexes, the kit’s proprietary Enzymatic Shearing Cocktail uses DNA digestion to release undissociated protein complexes from the DNA. Even histone complexes, which are not released from the DNA by traditional high-salt extraction methods, can be studied (Figure 2).
Figure 1: Co-Immunoprecipitation of a nuclear complex containing SRC-1 and ERα.
Figure 2: Detection of acetylated Histone H3.
Contents & Storage
Each Universal Magnetic Co-IP Kit provides sufficient reagents to make 25 nuclear and 25 whole-cell extractions, and then to perform 25 co-immunoprecipitations. Kits include PBS, Hypotonic Buffer, Whole-cell Lysis Buffer, Co-IP/Wash Buffer, Phosphatase Inhibitors, Protease Inhibitor Cocktail, Deacetylase Inhibitor, PMSF, EDTA, Digestion Buffer, Enzymatic Shearing Cocktail, Detergent, Protein G Magnetic Beads and a Bar Magnet for making a magnetic stand. The reagent storage conditions vary from room temperature to -20°C. Please download a manual for complete kit specifications and storage conditions. All reagents are guaranteed stable for 6 months when stored properly.
The following publications cite the use of and/or provide additional information about the Universal Magnetic Co-IP Kit:
- “PTEN regulates glioblastoma oncogenesis through chromatin-associated complexes of DAXX and histone H3.3.” by Benitez et al. (2017) Nature Communications 8(15223).