In Situ Hybridization Workflow

Due to their high affinity, high sensitivity, and low background, fluorescence- and enzyme-based detection reagents from Vector Laboratories are ideal for in situ hybridization (ISH) applications.

Step 1 Labeling

Labeling

Label your DNA or RNA probes and oligonucleotides using one-step direct methods, thiol-based indirect labeling, or specific 5' and 3' end tagging techniques.

Bioconjugation Reagents

Step 2 Characterization

Characterization

Confirm probe customization and label incorporation. For example, use the QuantTag Biotin Quantitation kit to determine the number of biotin molecules incorporated in each strand of nucleic acid.

Affinity Binding Matrices Biotin Quantitation

Step 3 Hybridization

Hybridization

Optimize the probe concentration and buffer conditions to ensure effective annealing to complementary sequences.

Slide Preparation

Step 4 Blocking

Blocking

Reduce or eliminate unwanted background (non-specific) staining on tissue sections and cell preparations using a blocking solution. Non-specific staining can result from endogenous enzyme or tissue elements, including endogenous enzyme activity, presence of Fc receptors, or interactions of detection reagents with tissue or cell proteins and other macromolecules. Choose a blocking solution based on the results of negative control sections.

Blocking Reagents

Step 5 Detection

Detection

Use fluorescent or enzyme-conjugated reagents that meet your sensitivity requirements and visualization preferences.

Fluorophore Conjugated Avidin/Streptavidin Enzyme Conjugated Avidin/Streptavidin Anti-Avidin/Streptavidin Reagents Antibodies to Tags & Labels Secondary Antibodies VECTASTAIN® ABC Kits (HRP) VECTASTAIN® ABC AP Kits (AP) ImmPRESS® Polymer Reagents for IHC

Step 6 Counterstain and Mount

Counterstain and Mount

Retain and preserve the specific target signal for short-term storage or longer term archiving.

Enzyme Substrates Counterstains and Special Stains Mounting Media