Standard Operating Procedure (SOP) for Tissue Decalcification
1.0 Purpose:
The purpose of this Standard Operating Procedure (SOP) is to provide guidelines for the decalcification of tissue samples, which involves the removal of calcium deposits, allowing for better tissue sectioning and histological analysis.
2.0 Scope:
This SOP applies to all personnel involved in tissue decalcification procedures within the laboratory.
3.0 Responsibilities:
The laboratory supervisor is responsible for ensuring that personnel are trained on this SOP and follow the procedures outlined.
The personnel performing the decalcification procedure are responsible for following the steps outlined in this SOP and maintaining the necessary documentation.
4.0 Materials and Equipment:
Formalin-fixed tissue samples
Decalcification solution (e.g., ethylenediaminetetraacetic acid (EDTA) or hydrochloric acid (HCl))
Specimen containers (e.g., plastic or glass vials)
Stirrer or shaker
Personal protective equipment (PPE), including gloves, lab coat, and safety goggles
Timer
pH indicator paper/strips (if using HCl decalcification solution)
Histology cassettes
5.0 Procedure:
Put on appropriate PPE, including gloves, lab coat, and safety goggles, before starting the decalcification procedure.
Label the specimen containers with unique identifiers to ensure proper sample identification throughout the process.
Fill the specimen containers with the decalcification solution, ensuring that there is enough volume to completely submerge the tissue samples.
If using EDTA decalcification solution:
a. Prepare the EDTA solution following the manufacturer's instructions.
b. Adjust the pH of the solution to the recommended range (typically between pH 7.0 and 7.4) using pH indicator paper/strips.
If using the HCl decalcification solution:
a. Prepare the HCl solution following the manufacturer's instructions.
b. Check the pH of the solution using pH indicator paper/strips. The recommended pH range is typically between 1.0 and 2.0.
Immerse the formalin-fixed tissue samples into the appropriate decalcification solution, ensuring that the samples are fully submerged.
Place the containers with the tissue samples and decalcification solution on a stirrer or shaker to facilitate agitation and enhance decalcification.
Set the timer according to the recommended decalcification time, which varies depending on the tissue type and size. Typical decalcification times range from a few hours to several days.
Monitor the progress of decalcification regularly by gently testing the tissue samples for firmness.
If using the EDTA decalcification solution, replace the solution with a fresh solution every 24 to 48 hours to maintain optimal decalcification conditions.
If using an HCl decalcification solution, monitor the pH of the solution periodically and adjust as necessary by adding additional acid or base to maintain the desired pH range.
Once decalcification is complete, remove the tissue samples from the decalcification solution and rinse them thoroughly with running water to remove any residual decalcification solution.
Place the decalcified tissue samples into histology cassettes for further processing and embedding.
Proceed with the next steps in the histological processing workflow, such as dehydration, clearing, and embedding, according to the laboratory's established protocols.
6.0 Documentation:
Record the date and time of starting and ending the decalcification process for each specimen.
Note the type of decalcification solution used and its concentration.
Document any observations or issues encountered during the decalcification process.
Maintain all records associated with the decalcification process as per laboratory guidelines.
Note: The decalcification protocol may vary depending on the specific laboratory requirements and the type of tissue being processed. It is essential to refer to any additional laboratory-specific guidelines or manufacturer instructions when performing tissue decalcification.
END OF THE DOCUMENT
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