Bio-Rad Clarity Western ECL Substrate User Manual

Clarity™ Western
ECL Substrate

Instruction Manual

Catalog # Description

170-5060 Clarity Western ECL Substrate, 200 ml
170-5061 Clarity Western ECL Substrate, 500 ml

Imaging

Table of Contents

Introduction 1

Quick Start Protocol 1

Background on Chemiluminescence 2

Antibody Incubation 2

Detailed Assay Procedure 5

Example Western Protocol 5

Immunodetection 5

Chemiluminescent Development 6

Troubleshooting 8

Ordering Information 11

Introduction

The Bio-Rad Clarity™ western ECL substrate is compatible with
any HRP-conjugate secondary detection reagent and ideal

for both digital and lm-based imaging. The Clarity substrate
provides excellent sensitivity with an extremely long signal
duration that allows re-imaging without loss of signal. In addition,
Clarity substrate is formulated to exhibit very low background
levels that yield exceptionally clear images. The combination of
bright, long signal, and low background makes Clarity substrate

the perfect choice for most blotting applications.

Quick Start Protocol

1. After immunodetection, keep the membrane moist in wash
buffer as you prepare the substrate mixture.

2. Mix substrate kit components in a 1:1 ratio. Prepare 0.1 ml

of solution/cm2 of membrane.

• For a mini-sized membrane

(7 x 8.5 cm), 7 ml of solution is sufcient

• For a midi-sized membrane

(8.5 x 13.5 cm), 12 ml of solution is sufcient

3. Place the membrane protein side up on a clear surface.

• Add substrate to the blot and incubate for 5 min

4. Image the membrane with a digital imager or by exposing to
X-ray lm.

Storage Conditions

Kit components are stable for at least one year at room

temperature. Freezing will not adversely affect kit performance
but do not expose to multiple freeze/thaw cycles.

Technical Support: 1-800-424-6723 • www.bio-rad.com 1