8.1 Centrifugation

8.1 Centrifugation

Quick Answer: Centrifuge by the lab’s RCF (× g), time, and temperature settings for each tube type—not by “whatever RPM the last shift used.” Balance every load, keep the lid closed until the rotor stops, give gel tubes full clotting time before spinning, prepare platelet-poor plasma for coag when required, and do not recentrifuge gel tubes after the barrier has formed unless a written exception exists. Broken tubes stay sealed until biohazard cleanup is ready.

Domain II (Specimen Preparation and Processing, 40–45% of the MLA exam) is the heaviest content area. Centrifugation sits at the center of that domain: serum and plasma chemistry, many immunoassays, and coagulation testing all depend on a clean, controlled spin. Poor technique creates hemolysis, fibrin strands, gel failures, and wrong platelet counts in plasma—errors that look like patient disease on the analyzer.

/practice/ascp-mlaPractice questions with detailed explanations

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RCF vs RPM: Speak the Language of Force

Centrifuge procedures are written in relative centrifugal force (RCF), expressed as × g (times gravity). Revolutions per minute (RPM) only describe how fast the rotor turns. The same RPM on two centrifuges with different rotor radii produces different g-forces.

Approximate conversion (use the formula on the instrument or manufacturer chart, not memory alone):

[ \text{RCF} = 1.118 \times 10^{-5} \times r(\text{cm}) \times (\text{RPM})^2 ]

Exam and bench habit: If the SOP says “1,000–1,300 × g for 10 minutes,” program g-force mode when the centrifuge supports it. If only RPM is available, convert using that rotor’s radius. Never copy RPM settings from a different brand or fixed-angle vs swing-bucket rotor without checking.

Typical patterns (always defer to local SOP and tube IFU):

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Balancing Loads

An unbalanced rotor vibrates, walks across the counter, damages bearings, and can shatter tubes.

Balance rules

1. Balance by mass, not only by “same number of tubes.” A full SST and a nearly empty tube are not a balanced pair.
2. Place tubes of similar volume and type opposite each other (180°).
3. Use water-filled balance tubes or manufacturer-approved ballast when odd numbers of patient tubes run.
4. Never balance a biohazard specimen against an open cup of water that can splash—use capped balance tubes.
5. For multi-position rotors, follow the diagram for 2-, 3-, 4-, and 6-tube layouts; empty positions must still leave the rotor symmetric.
6. Check adapters: a microtube in a full-size bucket needs the correct insert so it cannot tilt or eject.

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Gel-Barrier Tubes: Clot First, Then Spin

Serum separator tubes (gold/tiger SST) and many plasma separator tubes (PST) contain a thixotropic gel that migrates to form a barrier between cells and serum/plasma during centrifugation.

Clotting time before spin (serum tubes)

Clot-activator serum tubes need adequate upright clotting time before centrifugation—commonly about 30 minutes at room temperature for adult tubes, but follow the tube manufacturer and lab SOP (some rapid-clot tubes are shorter; cold samples clot slower).

Spinning too early:

• Leaves fibrin strands in the serum
• Causes incomplete barrier formation
• Clogs probes and invalidates results
• May force recollection

Plasma tubes (heparin PST) do not require clotting; they should be mixed gently at collection and spun per IFU timing (often sooner for STAT chemistry).

After the barrier sets: do not recentrifuge casually

Once the gel barrier has formed, recentrifugation is generally prohibited. Re-spinning can:

• Disrupt the barrier and remix cells into serum/plasma
• Alter analyte concentrations
• Introduce hemolysis or gel fragments

Do not recentrifuge a gel tube “to pack the gel better” or “to get a little more serum” after the barrier is set. If serum volume is QNS or fibrin remains, follow policy: may recollect, or in limited cases use a non-gel aliquot pathway directed by a technologist/supervisor—not MLA improvisation.

Some SOPs allow a single defined re-spin only under narrow conditions (e.g., incomplete first spin due to power failure before barrier migration)—those exceptions are written. Default exam answer: do not recentrifuge after barrier formation.

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Lid Closed and Aerosol Safety

• Close the centrifuge lid before starting; many instruments will not run with the lid open.
• Wait for complete stop before opening. Opening early can aerosolize contents if a tube fails.
• Prefer sealed rotors or safety cups for blood and body fluids when available; load and unload sealed carriers in a way that contains leaks.
• Position the centrifuge where vibration will not tip racks of open aliquots nearby.

Broken tube response

If you hear a crack, see liquid in the bowl, or open to a mess:

1. Do not immediately stick your face or hands into the open rotor.
2. Leave the lid closed for the time specified in the exposure-control plan (often ≥30 minutes) to allow aerosols to settle—follow your SOP exactly.
3. Don appropriate PPE (gown, gloves; face protection as required).
4. Open carefully; remove debris with forceps/tongs into a sharps or biohazard container as directed.
5. Disinfect the bowl, rotor, and adapters with the approved disinfectant and contact time.
6. Document the incident; notify the supervisor; determine which patient specimens were lost and need recollection.
7. Never salvage blood from glass shards into a new tube for testing.

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Platelet-Poor Plasma for Coagulation

Many coagulation assays (PT, aPTT, special coag) require platelet-poor plasma (PPP)—plasma with a very low residual platelet count (classically <10 × 10⁹/L, per method/CLSI-aligned lab goals).

Why it matters: residual platelets release phospholipids and factors that can shorten clotting times or interfere with lupus anticoagulant and other assays.

Double centrifugation is used when a single spin does not reliably produce PPP or when the special-coag SOP requires it. The MLA does not invent a second spin for convenience; follow the written method. Never harvest the buffy coat or cell layer into the plasma aliquot.

Hemolyzed, clotted, or underfilled citrate tubes are rejected before investing in PPP preparation.

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Practical Spin Workflow

1. Confirm orders, identifiers, tube types, and clotting status.
2. Group tubes by required RCF/time/temp (do not mix incompatible protocols in one “whatever” run if settings differ).
3. Balance the rotor; use sealed carriers when required.
4. Close lid; start run; do not walk away from a vibrating machine.
5. After stop, open safely; inspect for hemolysis, barrier quality, and volume.
6. Route serum/plasma to analyzers or aliquots without delay for labile analytes.
7. Log centrifuge maintenance issues (imbalance faults, broken glass, failed lid locks) per operations policy.

Common failure modes

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Key Takeaways

• Program RCF (× g) and time per SOP; convert RPM only with the correct rotor radius.
• Balance by mass, lid closed until stop, sealed rotors when available.
• Clot gel serum tubes fully before spinning; do not recentrifuge after the barrier sets without a written exception.
• Broken tubes: wait, PPE, disinfect, document, recollect—never salvage from shards.
• Coag needs platelet-poor plasma with validated single or double spins as protocol requires.