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Urinalysis & Body Fluids

Collection & Handling

We have just established the physiological foundation of the vaginal ecosystem, understanding it as a delicate, hormonally-driven balance. We must now turn to the most practical and, arguably, the most critical part of our investigation: the collection and handling of the vaginal secretion specimen

This is a unique pre-analytical challenge. Unlike a blood draw, the collection is often performed by a clinician at the point of care, and the most critical findings are highly perishable. The single most important organism we search for, Trichomonas vaginalis, is a fragile protozoan whose characteristic motility - the key to its identification - can be lost within minutes of collection if handled improperly

Therefore, “garbage in, garbage out” has never been more true. A flawless microscopic examination of a poorly collected or delayed sample is a futile exercise that will lead to a false-negative result and a failure to treat a patient’s infection. Our role as laboratory scientists is to be the experts who guide this process, providing the right tools and the right knowledge to ensure that the specimen we receive is a true and viable representation of the patient’s vaginal health

Purpose: The “Vaginitis Panel”

The collection is almost always aimed at diagnosing the cause of vaginitis, focusing on the “big three” common culprits: 1. Bacterial Vaginosis (BV) 2. Vulvovaginal Candidiasis (VVC, or “yeast infection”) 3. Trichomoniasis

A single collection event must yield enough sample for all the necessary point-of-care and laboratory tests

Collection Supplies: Right Tools for the Job

Using the correct supplies is non-negotiable

  • Speculum: A non-lubricated or water-lubricated speculum is used to visualize the vaginal walls and cervix. Gels and lubricants must be avoided as they are often bacteriostatic and can interfere with microscopy
  • Swabs: Dacron or rayon-tipped swabs on plastic shafts are required
    • CRITICAL: Do NOT use cotton-tipped swabs.: Cotton fibers are toxic to Trichomonas vaginalis and can interfere with certain molecular tests
    • Do NOT use wooden-shafted swabs.: The wood can contain resins that are inhibitory to some organisms and can interfere with PCR assays
  • Glass Microscope Slides: For immediate bedside preparation
  • pH Paper: A narrow-range (e.g., 3.0-5.5) pH paper is essential for accurate measurement
  • Reagents for Bedside Testing: 0.85% Saline (for wet mount), 10% Potassium Hydroxide (KOH) (for Whiff test and KOH prep)
  • Transport Media: Appropriate media for culture or Nucleic Acid Amplification Tests (NAATs) if being sent to the lab

Collection Procedure (Clinician-Performed)

This is a precise, multi-step process. The order is critical

  1. Step 1: Visualize and Assess pH (The FIRST Step)
    • After inserting the speculum, the clinician uses a dry swab to collect a sample of the vaginal fluid from the posterior fornix or the vaginal pool
    • The swab is then immediately rolled onto the pH paper
    • Rationale: The pH must be measured before any contact with saline or other reagents, as these will buffer the sample and give a falsely normal pH reading
  2. Step 2: Collect the Specimen
    • Using one or two new swabs, a generous sample of the vaginal discharge is collected from the vaginal pool. The swabs are rotated against the vaginal wall to collect epithelial cells
  3. Step 3: Immediate Slide Preparation (The MOST Time-Sensitive Step)
    • This is the core of the pre-analytical process.: The collected material must be immediately used to prepare two slides at the bedside
    • The Saline Wet Mount: One swab is immediately emulsified in a drop of room-temperature 0.85% saline on a slide, and a coverslip is applied. This slide is for the immediate microscopic search for motile trichomonads, clue cells, and WBCs
    • The KOH Prep: The second swab is emulsified in a drop of 10% KOH on another slide, and a coverslip is applied. This slide is for the microscopic search for yeast buds and pseudohyphae
  4. Step 4: The Amine (“Whiff”) Test
    • This is performed immediately after preparing the KOH prep.: The clinician brings the KOH slide close to their nose and sniffs it
    • Rationale: The 10% KOH is a strong base. It raises the pH of the sample dramatically, which volatilizes the amines (putrescine, cadaverine) produced by the anaerobic bacteria in Bacterial Vaginosis, releasing a characteristic “fishy” odor. A positive Whiff test is a classic sign of BV
  5. Step 5: Inoculate Transport Systems
    • If additional testing (e.g., yeast culture, NAATs for Trichomonas or a BV panel) is required, a third swab is used to inoculate the specific transport medium provided by the laboratory

Specialized Collections

  • Group B Streptococcus (GBS) Screening: This is a crucial screening test in late-stage pregnancy. The collection is different. It requires a vagino-rectal swab the swab is first inserted into the lower vagina and then inserted into the rectum. The swab is then placed into a specific transport medium (like Amies or Stuart’s) to preserve the bacteria for culture
  • Nucleic Acid Amplification Tests (NAATs): These are highly sensitive molecular tests for organisms like Trichomonas, Candida species, and the bacteria associated with BV. They are also the gold standard for STIs like Chlamydia and Gonorrhea
    • These tests require a specific, lab-provided collection kit with a designated swab and transport medium
    • Self-Collection: A major advantage of NAATs is that they are highly effective with patient-collected vaginal swabs. This improves patient comfort and access to testing. The patient is given a kit with clear instructions on how to insert the swab into the vagina, rotate it, and place it in the transport tube

Transport & Handling: Race Against Viability

STAT Priority: Trichomonas Motility

  • Urgency: The saline wet mount for Trichomonas detection is a STAT test. It should ideally be examined by an experienced microscopist within minutes of collection at the point of care
  • Time Limit: If sent to the lab, the unpreserved swab or prepared wet mount must be analyzed within 1-2 hours maximum. After this, the characteristic jerky motility is lost, and the organism becomes very difficult to differentiate from a white blood cell, leading to false negatives
  • Temperature: The specimen must be maintained and transported at ROOM TEMPERATURE.
    • Do NOT refrigerate!: Refrigeration will rapidly kill Trichomonas vaginalis and render it non-motile

General Handling

  • Slides should be protected from drying out. A “moisture chamber” (a petri dish with a moist paper towel) can be used for transport to the lab
  • Swabs for culture or NAATs should be transported according to the specific manufacturer’s or laboratory’s instructions, but this is typically at room temperature

Specimen Rejection Criteria

The lab must act as the final quality gatekeeper. A specimen should be rejected if:

  • Improper Swab: A cotton or wooden-shafted swab is received for Trichomonas or NAAT testing
  • Dry Swab: A swab for wet mount or culture arrives completely dry
  • Excessive Delay: A sample for Trichomonas motility testing is received more than 2 hours after collection
  • Improper Temperature: A sample for Trichomonas is received on ice
  • Unlabeled or Mislabeled: As with any specimen, this is an absolute cause for rejection

Conclusion

The pre-analytical phase for vaginal secretions is a model of the crucial partnership between the clinic and the laboratory. Success hinges on using the correct tools, following a precise order of operations, and, above all, respecting the extreme fragility of our most important diagnostic target: the motile trichomonad. By mastering these principles, we ensure that the microscopic and molecular tests we perform provide a true and accurate assessment of the vaginal ecosystem, leading to the correct diagnosis and effective treatment for the patient