01_Specimen Collection, Transport and Processing

• differential diagnosis


  Identify the possible disease to narrow down the

  tests needed or recommended
• Pre-analytical phase 


  most critical phase and done prior to submission to the laboratory
• The Rule of Thumb in pre-analytical phase:

  Garbage in = garbage out
• Usual Lab acitivities? (BVPSM)

  • Bacterial Culture
  • Virus Culture
  • PCR Testing or Molecular Testing
  • Serology
  • Microscopy
• optimum VTM is - does not require refrigeration
• optimum timing of:

  • respiratory swab specimens - 3-7 days
  • Whole blood, CSF - as soon as possible
• sample types (6-SWCSUA)

  • Stool,
  • Whole blood,
  • CSF,
  • Sputum,
  • Urine,
  • Autopsy tissues
• Why you might not be able to use VTM for bacterial specimens?

  VTM will have a little antibiotic which would bring a false-negative result towards bacterial culture. If antibiotics are already given, an antimicrobial solution is applied to neutralize the antibiotic effect
• Refrigeration of serology specimen:

  • 4-8 °C for 24-48 hours
  • -20 °C for longer periods
• Adequate Volumes

  • PCR tests - <1.0 mL
  • Culture tests - more
  • VTM - >3.0 mL


  Some specimen types should NOT be placed in transport media. sometimes undiluted is optimal
• Tourniquet - strip of cloth or rubber used in applying pressure to limit or prevent blood flow
• blood smears are fixed with?

  Methanol
• Handling & Transport: Blood Smear

  • Transport slides within 24 hours
  • Do not refrigerate as this can alter cell morphology of the blood
• Collection: Blood for collection

  • Infants: 0.5 - 2 mL
  • Children: 2 - 5 mL
  • Adults: 5 - 10 mL
• Collection of blood for culture requires aseptic technique and within 10 minutes of fever.

  
  

  If suspect bacterial endocarditis: 3 sets of blood culture
• Handling and Transport: Blood for cultures

  • Collect into bottles with infusion broth
  • Change the needle to inoculate the broth
  • Transport upright with cushion to prevent hemolysis
  • Wrap tubes with absorbent cotton or gauze
  • Travel at ambient temperature within 24 hours
  • Store at 4 °C if more than 24 hours
• Collection of Blood serum

  • Let blood clot for 30 minutes at ambient temperature
  • Blood will clot within a few minutes
  • Glass activates clotting faster
  • Purple cap contains a commonly used EDTA preservative that prevents clotting of blood, and by centrifuging, it will result in plasma
  • Red cap means there is no preservative and is empty
• Handling of blood serum:

  • Place at 4-8 °C for clot retraction for at least 1-2 hours
  • Centrifuge at 1,500 RPM for 5-10 minutes
  • Separate serum from the clot immediately
• Transporting Blood Serum:

  • 4-8 °C if transport lasts less than 10 days
  • Freeze at -20 °C if storage for weeks or months - can last for a year in storage
  • repeated freeze-thaw: destroys IgM
  • freezing unseperated blood - causes hemolysis
• Patient Factors affecting Hemolysis

  • Metabolic Disorders
  • Chemical Agents
  • Physical Agents
  • Infectious Agents
• Phlebotomy Factors affecting hemolysis

  • Catheter Collections
  • Capillary Collection
  • Needle Gauge
  • Location of Venipuncture
  • Antiseptic Used
  • Tourniquet time
  • Traumatic Draw
  • Tube type
  • No or Vigorous mixing
  • Syring Transfer
• Processing Factors that affect homolysis

  • Time between collection and centrifugation
  • Temperature and Speed of centrifuge
  • Poor separator barrier Integrity
  • Recentrifugation
• Collection: CSF Samples

  • lumbar puncture is between Lumbar spine: L3, L4, and L5
  • under aseptic conditions
• Handling and Transportation of CSF (Bacteria):

  • in trans-isolate medium
  • Pre-warmed to 25-37 °C before inoculation to avoid temperature shock
  • or transport to ambient temperatures
• Handling and Transportation of CSF (Virus):

  • 4-8 °C if up to 48 hours,
  • -70 °C for a longer duration
• Urine colors

  • Colorless
  • Pale yellow
  • Dark yellow/Amber/Orange
  • Pink/Red
  • Brown
  • Port Wine
  • Green
  • Blue-Green
  • Blue
  • Black
  • Milky
• Urine clarity

  • Clear - No visible particulates; transparent
  • Hazy -Few particulates print easily seen through urine
  • Cloudy - Many particulates; print blurred through urine
  • Turbid - Print cannot be seen through urine
  • Milky - May precipitate or be clotted (like condensed milk)
• Urine Odor

  • Faint aromatic odor - NormalAmmoniacal - Characteristic of Bacterial pathogens breaking down urea
  • Foul odor - Old specimen
  • Sweet - Presence of ketones
  • Maple syrup - Maple Syrup Urine Disease (MSUD)
  • Mercaptan - After eating asparagus
  • Rotting fish - Trimethylaminuria
  • Rancid butter - Tyrosinemia
  • Cabbage - Methionine malabsorption
  • Mousy - Phenylketonuria (PKU)
  • Sweaty-feet - Isovaleric acidemia
• Collection of urine

  • random specimen - urine is collected at an unspecified time

  Used for:

  • Urinalysis testing
  • Measurement of specific gravity
  • pH
  • Glucose levels
• Urine dipstick kit - Basic qualitative diagnostic tool that tests for several parameters (10) to determine pathological changes in the urine sample
• Technique used for culture and Sensitivity?

  Midstrem Specimen
• Urine Culture Test - performed to grow and identify organisms, mainly bacteria and fungi, that may cause Urinary Tract Infections (UTI)
• Antibiotic Sensitivity/Susceptibility Test - helps to select an appropriate antibiotic that is effective against specific types of bacteria or fungi causing any infection
• Collection methods of sterile urine

  • Suprapubic Aspiration - using a syringe, urine is collected directly from the bladder
  • Indwelling Catheter - Aseptic, collects urine by attaching to a drainage bag.
• Timed urine specimen is collected in 2-72 hour intervals; commonly every 24 hours
• Collection: Stool Samples

  • Disinfectant residues may kill microorganism
  • avoid moisture as there are microorganisms in water
• • Advantages of rectal swabs - Convenient for for children
  • Disadvantages - No macroscopic test, less, not for viral detection
• Stool Samples for viruses

  • Time: within 48 hours
  • Size: 5-10 mL
  • Method: unmixed, dry, sterile
• Stool Samples for Bacteria

  • Time: active phase
  • Size: fresh, two swabs
  • Method: Cary-Blair medium, no tranpost medium (Ag/PCR)
• Stool Samples for Parasites

  • Time: ASAP
  • Size: 3 x 5-10 mL fresh stool
  • Method: 10% formalin or polyvinyl chloride (3:1); Unpreserved samples for Ag detection and PCR