otsB Antibody, HRP conjugated

Target Biology and Function

The otsB gene encodes trehalose-6-phosphate phosphatase, a critical enzyme in trehalose biosynthesis. Trehalose acts as a stress protectant in E. coli, enabling survival under osmotic stress, heat, and desiccation . The antibody binds specifically to the otsB protein, enabling quantification of its expression levels under experimental conditions.

Primary Use

• ELISA: The HRP conjugation allows chromogenic or chemiluminescent detection of otsB in E. coli lysates or purified samples .

Potential Extended Uses

While current documentation specifies ELISA, HRP-conjugated antibodies are broadly used in:

• Western Blotting: For protein size validation (requires validation for otsB) .

• Immunohistochemistry (IHC): Localization of otsB in bacterial colonies (theoretical, pending protocol optimization) .

Conjugation Methodology

The HRP conjugation process typically involves:

1. Antibody Modification: Introduction of reactive groups (e.g., thiols) via crosslinkers like Sulfo-SMCC .

2. HRP Activation: Maleimide-activated HRP is reacted with the modified antibody to form stable thioether bonds .

3. Purification: Removal of unreacted HRP or antibodies via size-exclusion chromatography .

Performance Enhancements

• Lyophilization of activated HRP improves conjugation efficiency and stability, as demonstrated in analogous studies .

• Site-specific conjugation (e.g., heavy-chain targeting) ensures consistent labeling ratios (1–2 HRP molecules per antibody), enhancing signal reproducibility .

HRP Conjugation Kits

Validation and Quality Control

• UV Spectroscopy: Confirms HRP-antibody conjugation via peak shifts (e.g., HRP peak at 430 nm) .

• SDS-PAGE: Validates molecular weight shifts due to HRP attachment .

• Functional ELISA: Demonstrates sensitivity down to 1:5,000 dilutions in optimized protocols .