iMMUNOFLUORESCENCE ASSAY LAB CHECKLIST

Ensure all reagents and materials are available and properly labeled.

Check expiration dates on all reagents.

Prepare and label all required buffers and solutions (e.g., PBS, fixation buffer).

Gather necessary equipment (e.g., microscope, pipettes, centrifuge).

Review the experimental protocol and objectives.

Collect tissue or cell samples according to the established protocol.

Fix samples using an appropriate fixative (e.g., formaldehyde, methanol).

Permeabilize samples if required (e.g., using Triton X-100).

Block non-specific binding sites with a blocking solution.

Dilute primary antibodies according to the manufacturer’s instructions.

Prepare secondary antibodies if necessary, ensuring they are compatible with the primary antibodies.

Include appropriate controls (e.g., isotype controls, no primary control).

Incubate samples with primary antibodies for the recommended time and temperature.

Wash samples thoroughly to remove unbound antibodies.

Incubate with secondary antibodies, if used, for the required duration.

Perform additional washes to eliminate unbound secondary antibodies.

Mount samples using an appropriate mounting medium containing anti-fade agents.

Ensure samples are coverslipped correctly to avoid air bubbles.

Set up the fluorescence microscope and adjust settings for optimal imaging.

Capture images of the samples based on the experimental design.

Analyze images using appropriate software.

Quantify fluorescence intensity if necessary.

Compare results with controls and replicates.

Document findings and prepare reports.

Dispose of all hazardous waste according to safety protocols.

Clean all equipment and workspace thoroughly.

Store any remaining reagents properly for future use.

Review and organize lab notebooks or digital records for the experiment.

Ensure proper personal protective equipment (PPE) is worn throughout the experiment.

Follow all institutional biosafety guidelines.

Conduct a post-experiment review and note any issues encountered for future reference.