Donkey Anti-Goat IgG/Cy5.5

The Donkey Anti-Goat IgG/Cy5.5 antibody is a valuable tool for various immunofluorescence (IF) applications. Below are the protocols for the validated application.

For IF, the following steps are recommended:

Reagents or buffers: Phosphate-buffered saline (PBS), 4% paraformaldehyde (PFA) for fixation, 0.1% Triton X-100 for permeabilization, and a blocking buffer (e.g., 5% normal donkey serum).

Sample preparation: Fix cells or tissue sections with 4% PFA, followed by permeabilization with 0.1% Triton X-100. Block with 5% normal donkey serum to reduce non-specific binding.

Antibody incubation: Dilute the Donkey Anti-Goat IgG/Cy5.5 antibody to 1:50-500 in the blocking buffer and incubate with the sample for 1-2 hours at room temperature or overnight at 4°C.

Washing or detection: Wash the sample thoroughly with PBS, then proceed with fluorescence imaging. The Cy5.5 fluorophore has an excitation maximum at approximately 675 nm and an emission maximum at approximately 694 nm.

Technical notes: Optimize the antibody dilution and incubation time based on the specific application and sample type. Use appropriate controls, such as omitting the primary antibody, to verify the specificity of the staining.

Use assay conditions validated for IF, apply Donkey Anti-Goat IgG/Cy5.5 at the recommended working dilution, include proper positive and negative controls, perform thorough wash steps, and document incubation time, temperature, and detection conditions for reproducible performance. Recommended dilution: IF=1:50-500.