An immuno-reaction enhancing solution for Western Blotting and ELISA

IMMUNO SHOT (Reagent 1 & Reagent 2)


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Background

IMMUNO SHOT is an enhancer of antigen-antibody reaction. In Western blotting and ELISA, researchers often experiences weak signal or high background. IMMUNO SHOT improves these problems by just using it as antibody diluents. Due to the principle of working mechanism, IMMUNO SHOT can be used in many assay systems that use antigen-antibody reaction.

How IMMUNO SHOT Works
IMMUNO SHOT contains a polymer which, by changing the physicochemical properties of antigen and antibody, enhances the mutual accessibility, and facilitate the specific reaction. The other ingredient, protein, reduces non-specific binding of antibody. Thus, IMMUNO SHOT enhances the antigen-antibody reaction while reduces background.

Application

œIMMUNO SHOT is consisted of Reagent 1 and Reagent 2.Use Reagent 1 for the dilution of the 1st antibody.Use Reagent 2 for the dilution of the 2nd antibody.No change is required for the other assay protocol.

œIn some assay systems, only one antibody is used.For example some ELISA uses only one enzyme-conjugated antibody.In such case, try using Reagent 2 for the antibody dilution.In some cases, however, Reagent 1 gives better result.

œIMMUNO SHOT has been successfully used in Western blotting, antibody sandwich ELISA with either the 1st or 2nd antibody-labeled type, antigen sandwich ELISA

 

<Western Blotting>
Protocol of Western Blotting (WB)

<ELISA>
Protocol of ELISA

<Troble shooting>
Trouble shooting

   

Feature and Advantages

1.@High signal with low background
IMMUNO SHOT enhances the antigen-antibody reaction. Comparing with the method using detergent-containing buffer, several to over 10-fold stronger signal can be obtained while the background level is low. Thus, you can get much higher S/N ratio than usual method.

2.@Effective for saving antibody usage and time of reaction time
Because higher signal can be obtained using IMMUNO SHOT, you can reduce the amount of antibody used and the time required for reactions.

3.@Can be used for many reactions
IMMUNO SHOT can be used not only for Western blotting and ELISA, but also for other assay systems using antigen-antibody reactions. In addition, IMMUNO SHOT does not affect activities of HRP (horse radish peroxidase) or AP (alkaline phosphatase), and can be used for assay systems using these enzymes.

4.@Easy to use
IMMUNO SHOT is formulated as to Ready to Use. Just exchange your dilution buffer of antibodies to the solutions of IMMUNO SHOT.

Application Exapmple

Protein detectinon

Example - 1  Detection of RhoG Protein
MDA-MB-231Cell Lysate
Primary antibody (Diluted by Reagent 1):Anti-RhoG, mouse monoclonal, 1/500
Secondary antibody(Reagent 2 dilution):Anti-mouse IgG, goat polyclonal HRP,1/3000
DetectionF@ECL, Fuji LAS-3000

 

Example - 2  Detection of diphospho-ERK (MAPK)@Protein
S2@cell lysate
Primary antibody( Diluted by Reagent 1):Anti-diphospho-ERK, mouse monoclonal, 1/5000
Secondary antibody( Diluted by Reagent 2):Anti-mouse IgG, goat polyclonal HRP,1/10000
Detection:@ECL, Film for ECL, Light exposure 15 minutes

 

ELISA

Example - 1
ELIS Aassay: Antibody for HBsAg S Antigen activation detection, Sandwich ELISA
Result: On the case using both primary antibody and seondary antibody, and the case using only primary antibody, IMMUNO SHOT shows much stronger signals comparing to the case using PBS-T
IMMUNO SHOT also shows superior strangh of signal compering to A company reagent.

 

Detection using primary antibody and secondary antibody



Detection using only primary antibody



 

 

 

Western Blotting

Example - 1

Sample: HepG2 Cell Lysate that express Luciferase expression
How to test: Transfer Lysate to PVDF membrane after SDS-PAGE, conbine primary (Anti-Luciferase IgG, goat polyclonal, 1/1000), and conbine secondary antibody (Anti-Goat IgG , donkey HRP-labeled, 1/20000). Protein band are detected by 5min exposure with ChemiDoc XRS (Bio-Rad) from chemiluminescence@caused by HR activation with ECL solution.  
For the dilution of primary antibody and secondary antibody, IMMUNO SHOT Reagent 1AReagent 2, and Competitore A enhancer (Solution1 and Solution") are used.  In NT case, TBS-T (0.1 % Tween-20) are used for dilution of both primary antobody and secondary antibody.
Result: IMMUNO SHOT shows much stronger signals compering to the case usignf TBS-T.
IMMUNO SHOT  shows equal (or more) strenghth comparing to the case using competitor A reagent.

Example - 2
Sample: A431Cell Lysate
How to testF After transfer Lysate to SDS-PAGE, combine primary antibody(Anti-human EGFR IgG,@mouse monoclonal), and combine secondary antibody(Anti-mouse IgG, rabbit polyclonal HRP-labeled, 1/10000) . The protein band are detected by 10 min exposure with ChemiDoc XRS(Bio-Rad), from chemiluminescence@caused by HR activation with ECL solution. For the dilution of primary antibody and secondary antibody, IMMUNO SHOT Reagent 1AReagent 2, and Competitore A enhancer (Solution1 and Solution")  are used. In NT case, TBS-T (0.1 % Tween-20) are used for both primary antobody and secondary antibody.
Result: IMMUNO SHOT shows much stronger signals compering to the case usign TBS-T.

Product List

Product Name Cat# Quantity Price

IMMUNO SHOT Reagent 1&2 DataSheet ,

CSR-IS-012-100

1 KIT ¥9,800
$131

IMMUNO SHOT Reagent 1&2 DataSheet ,

CSR-IS-012-250

1 KIT ¥18,000
$240

IMMUNO SHOT Reagent 1 DataSheet ,

CSR-IS-001-250

250 ML ¥9,500
$127

IMMUNO SHOT Reagent 2 DataSheet ,

CSR-IS-002-250

250 ML ¥9,500
$127

To be used for research only. DO NOT use for human gene therapy or clinical diagnosis.



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