Antigen-Antibody Binding Enhancer Kit                 Cat# W-9812              Price: $79/Set

 Benefits:

1. Money Saving:  Usually  with  30-50% less antibody we get good results, please see the Figs below.

2, Time saving: 30 minutes to 90 minutes protocol available.

 Kit Content:

  1,  20 x Binding Buffer(20xBB) 10 ml.

  2,  100 x Binding Enhancer (100xBE) 2 ml.

Note: Milk powder / BSA Powder is not include in the kit.

 Calculation:   Make 10 ml 1x Binding Buffer (1xBB) --  mix 20xBB 0.5 ml,  100xBE 0.1 ml,  Distilled Water (ml) 9.4 ml and 

 Milk / BSA  0.5g. please make sure   Milk powder / BSA is dissolved completely.

                             

                              Quick  Protocol (For the primary & Second antibody to be used together)

1. The ready Western membrane is blocked in wash buffer contained 5% Milk/BAS, to be shaking at RT for 2 minutes 

   (5-10 min. are allowed).

2. Put the blocked Membrane in the 1x BB contained the  primary and HRP conjugated second antibody  together, shake at RT

   for 20 minutes, (30-60 minutes is allowed if protein has low expression)

3. Rinse the membrane with wash buffer for 4-6 times and continue to wash it with wash buffer at RT, shake for 2 minutes by 

   4 times.

4. The membrane has been ready to be developed with ECL.

 

                           One Hour Protocol (For the primary & Second antibody to be used separately)

1.The ready Western membrane is blocked in wash buffer contained 5% Milk (or BAS),  shake at RT for 2-4 minutes 

(5-10 min. are allowed).

2. Put the blocked Membrane in the 1xBB contained acceptable primary antibody, to be shaking at RT for 20 minutes, 30-60

 minutes is allowed if there is a week expression.

3. Rinse the membrane with wash buffer for 4-6 times and continue to be washed with wash buffer at RT, shake for 2 minutes 

 by 4 times.

4. Put the membrane in 1xBB with acceptable second antibody; shake at RT for 20 minutes.

5. Rinse the membrane with wash buffer for 4-6 times and continue to wash it with wash buffer at RT, shake for 2 minutes  by

 4 times.

6. The membrane has been ready to be developed with ECL.

 Reference

  1.    2.    3.

                                                                                                           

Fig.1 The nitrocellulose membrane. Mouse tissue protein.  With anti-actin, 1:30,000.

Fig.2 Human cancer tissue (B, Cell line only) protein. The Phospho-P44/42 (thr202/tyr204); MAP Kinase antibodyl, 1:1000).

Fig.3  , Human cancer cell line protein, Anti-P-FADD (ser 191)  1:1000

 

4.     5.   

Fig.4.the proteins were from the human cell line transfected with anti-Cyclin D1, 0.83 mg/ml.

Fig. 5, Human cell line protein, antibody-NFêB p50, 0.66ug/ml.

 

 

 

 

 

 

          

 

 

 

 

                                    This product is distributed for laboratory research only.Caution: Not for diagnostic use.