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Index / Geba / Antibody Stripping Buffer / Product Detail : ST010 Antibody Stripping Buffer
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Jul
28th

#ST010 Antibody Stripping Buffer

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  Price : 75   EUR
86   USD
58   GBP
319   Zloty
10162   JPY
585   NOK
627   SEK
85   CHF

Product name : Antibody Stripping Buffer

Catalog number : ST010

Quantity: 1

Availability: Yes

Supplier name : Geba

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More Details about

Cat. No.  Description
ST010  Immunodetection removing buffer, 500 ml 
ST013  Immunodetection removing buffer, 30 ml 
Storage and Shipping Instruction:  Shipped at ambient temperature. Upon arrival,
store at 4oC. Stable at room temperature or 4oC for 1 year.
IMPORTANT: Allow the buffer to warm to room temperature before use.
  
This product is guaranteed for one year from date of purchase when properly handled and stored.


Introduction
Western Blot is widely used to detect and compare proteins from a complex mixture utilizing antibody detection on a membrane. Chemiluminescence has become an easy and sensitive method of detection compared to other methods of analysis. Because of the nature of chemiluminescence detection, it is possible to  reprobe the separated protein mixture on the membrane. Conventionally, Western blots have been stripped using extremely harsh conditions that  may alter the antigen for subsequent immunoprobing. The Gene Bio-Application  Stripping Buffer is a novel formula
providing a gentle method of removing  primary and secondary antibodies from membranes that allows reprobing the same membrane several times.   


Protocol
IMPORTANT: Optimization of incubation time is essential for best results.
IMPORTANT: If the blot cannot  be stripped immediately after chemiluminescence detection, the  blot can be stored in PBS or
TBST at 4o C until the stripping procedure is performed. 
   
1.  Place the blot to be stripped in the Stripping Buffer. Top up Stripping
Buffer until the blot is completely immersed.

2. Incubate the blot in the Striping buffer for 5-15 min at room temperature
under vigorous shaking.
IMPORTANT: In general, higher  affinity antibodies or large quantities of detected protein will require longer incubation time for stripping. However, each half-minute is critical to achieve optimum incubation time.
 
3. Empty the Stripping Buffer. 
4. To wash, add 300 ml of dH2O and shake vigorously for 5 min.
Repeat Step 4 five more times.
Verification of complete removal of the HRP label: After Step
5 incubate the membrane with  fresh chemiluminescence reagent and expose to film. If no signal is detected after 5 min exposure, the HRP conjugate has been  successfully removed from the
antigen or primary antibody.
Verification of complete removal of primary label antibody:
After Step 5 incubate the membrane with the HRP-labeled secondary antibody, followed by a wash in wash buffer. Incubate the membrane with fresh chemiluminescence reagent and expose
to film. If no signal is detected after 5 min exposure, the primary antibody has been successfully removed from the antigen.
IMPORTANT:  Analysis of successful removal of immunoprobes is recommended to prevent removal of the antigen or the unsuccessful removal of the antibodies. 
6. If signal is detected in the two experiments described above, place the blot again in Stripping Buffer for additional 5-15 min.
7. After it has been determined that the membrane is free of the immunodetection reagents, a second immunoprobe can take place.
Start the second immunoprobe with reblocking of the blot. 
IMPORTANT: The blot can be stripped up to 5 times.
However, longer exposure times or more sensitive chemiluminescence substrate is  needed. Actually, re-probing may result in a decrease in signal if the antigen is unstable.
Analysis of the individual system is required.  

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