**Specimen Preparation

Specimen preparation is a crucial step in the pre-analytical phase of laboratory testing. It involves preparing the collected blood samples for analysis. This includes centrifugation to separate the cellular components from the liquid portion (serum or plasma), visual inspection, and aliquoting the sample into smaller portions for different tests. Errors in this stage can lead to inaccurate results, potentially impacting patient care. The goal is to obtain a high-quality sample that is representative of the patient's condition.

Centrifugation is the process of using centrifugal force to separate substances of different densities. In phlebotomy, it's primarily used to separate blood cells from serum or plasma.

Key principles:

• Speed (RPM - Revolutions Per Minute) and Time: The speed and duration of centrifugation depend on the type of specimen and the desired separation. Follow the laboratory's Standard Operating Procedures (SOPs) for specific guidelines. Generally, serum and plasma samples require centrifugation at a specified speed (e.g., 1500-2500 RPM) for a specific time (e.g., 10-15 minutes). Whole blood samples typically require a lower speed and shorter duration if centrifugation is even necessary.
• Balance: Proper balancing of the centrifuge tubes is essential to prevent damage to the centrifuge and ensure even separation. Always use tubes of equal weight, and arrange them in the centrifuge rotor in a balanced manner (opposite each other).
• Temperature: Some analytes are sensitive to temperature. Refrigerated centrifuges are used to maintain sample integrity and prevent degradation of certain components, such as some coagulation factors. Check the requirements for each specific test you're performing.

Example: A serum separator tube (SST) containing a clot activator and gel separator, once centrifuged, will result in the blood cells being pushed to the bottom of the tube by centrifugal force, and a layer of gel separating the serum (the liquid portion, devoid of cells) from the clot.

Serum: Serum is the liquid portion of the blood that remains after the blood has clotted. It does not contain clotting factors (like fibrinogen) because they are used up during the clotting process.

• Process: Allow the blood in a red-top tube (or other tubes with clot activators) to clot for 30-60 minutes before centrifugation. After clotting, centrifuge the tube to obtain the serum. Carefully pipette the serum away from the clot.

Plasma: Plasma is the liquid portion of the blood that still contains clotting factors. It is obtained by centrifuging blood collected in tubes containing anticoagulants (e.g., EDTA, heparin, citrate).

• Process: Immediately after collection, centrifuge the blood to separate the plasma from the blood cells. Plasma samples are often used for STAT (immediate) testing because they don't require clotting time.

Whole Blood: Whole blood is used for certain tests that require all the blood components, such as complete blood counts (CBCs).

• Process: Usually, no centrifugation is required for whole blood tests. The sample is directly analyzed after proper mixing (gently inverting the tube several times to ensure the anticoagulant, if present, is well-mixed). Centrifugation may be performed for some specialized tests, but the laboratory SOPs must be consulted.

Aliquoting involves transferring a portion of the original sample into smaller tubes or containers. This is done for several reasons:

• To run multiple tests from a single sample.
• To store excess sample for potential repeat testing or reflex testing.
• To send samples to a reference laboratory.

Procedure:

1. Labeling: Ensure all aliquot tubes are properly labeled with the patient's information, including patient name, medical record number, and date and time of collection. This is CRUCIAL to prevent errors.
2. Equipment: Gather appropriate pipettes (manual or automated), aliquot tubes, and any other necessary materials (e.g., gloves, absorbent pads).
3. Mixing: Gently mix the original sample (e.g., by inverting the tube) to ensure the components are evenly distributed.
4. Dispensing: Carefully pipette the required volume of the sample into the labeled aliquot tubes. Avoid splashing and contaminating the sample.
5. Storage: Store the aliquot tubes according to the laboratory's guidelines for temperature and duration (e.g., refrigerated, frozen). Proper storage protects sample integrity. Specific storage conditions will be test-dependent (e.g., some analytes are stable at room temperature for a short time, while others must be frozen immediately).

Important Considerations:

• Use the correct pipette tip for the sample type and volume.
• Avoid cross-contamination. Use fresh pipette tips for each sample and each aliquot.
• Do not overfill or underfill aliquot tubes.
• Close the aliquot tubes tightly to prevent leakage or evaporation.

Several errors during specimen preparation can affect test results:

• Hemolysis: Rupture of red blood cells, releasing hemoglobin into the serum or plasma. Caused by improper collection, rough handling, or incorrect centrifugation.
• Clotting in Anticoagulated Tubes: Failure to properly mix blood with anticoagulant, leading to clot formation. Always invert tubes gently but thoroughly after collection.
• Incorrect Centrifugation: Using incorrect speed or time can lead to incomplete separation or damage to the sample components.
• Improper Storage: Storing samples at the wrong temperature or for too long can degrade analytes.
• Contamination: Using contaminated equipment or transferring samples incorrectly can introduce errors.
• Incorrect Aliquot Volumes: Using insufficient or excessive sample volumes can compromise test accuracy. Read the SOPs carefully.