Key facts
- Brand
- Agilent
- Model
- P200
- Industry
- Industrial
- Documents
- 5
- Languages
- EN
- Last added
- 2012-07-05
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Agilent P200 ScreenTape Suppression of Antibody Reduction Artifacts Technical Overview
This document provides a technical overview regarding the analysis of monoclonal antibodies (mAbs). It addresses the issue where certain IgG subtypes exhibit higher than expected half-antibody representation under denaturing, non-reducing conditions due to cryptic cysteins exposed by dodecyl sulfate.
The text details an experiment using N-ethylmaleimide (NEM) prior to SDS-PAGE analysis. The results show that treating labile IgGs with NEM reduces the formation of these denaturing artifacts on P200 ScreenTape under non-reducing conditions, offering a more accurate method for mAb quantification compared to traditional methods.
Agilent P200 specifications
| Staining reaction optimal pH range | pH 8-10 |
|---|---|
| P200 pH buffer pH | pH 9 |
| Urea compatibility limit | 8M |
| Guanidine compatibility limit | 600 mM |
| Salt concentration limit for migration | 500 mM |
| Triton X-100 compatibility limit | 1% |
| Acetonitrile compatibility limit | 20% |
| Methanol compatibility limit | 20% |
| Ethanol compatibility limit | 20% |
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The shortest path to a working setup, usually only a few pages.
How to use the product for one specific task, with worked examples.
Questions answered in these manuals
How does NEM treatment affect antibody analysis on P200 ScreenTape?
Treatment with N-ethylmaleimide (NEM) prior to SDS-PAGE analysis reduces the appearance of half antibodies by alkylating free cystein residues, preventing auto-reduction.
From: Agilent P200 ScreenTape Suppression of Antibody Reduction Artifacts Technical Overview
What is the recommended pH for the NEM reaction?
The reactions should be carried out at a neutral pH because elevated pH can cause NEM to react with lysine residues and produce unexpected results.
From: Agilent P200 ScreenTape Suppression of Antibody Reduction Artifacts Technical Overview
What is the optimal pH range for the staining reaction?
The staining reaction occurs optimally between pH 8-10 as the amine group needs to be deprotonated to be available to the dye.
Can guanidine be used in sample preparation?
Guanidine has low compatibility at 600 mM because it causes SDS precipitation. It is recommended to dilute guanidine to below 600 mM or use urea instead.
Can P200 ScreenTape analyze samples under different conditions?
Yes, it allows analysis of samples under both reducing and non-reducing conditions.
From: Agilent P200 ScreenTape Application Note for Antibody Testing
How many samples can be analyzed without manual intervention?
Integrity and purity analysis of up to 16 mAb preparations can be performed in less than one minute per sample.
From: Agilent P200 ScreenTape Application Note for Antibody Testing