Specimen Collection pg 6-10

first 7-14 days

Diagnostic yield is highest when samples are obtained within the __ of symptom onset, when viral shedding is at its peak. Sensitivity decreases significantly with delayed collection.

Personal Protective Equipment (PPE)

N95 respirator, disposable gown, gloves, and protective eyewear (goggles or face shield).

Positioning

Collectors should stand slightly to the side of the patient to minimize exposure during coughing or sneezing.

Collection environment:

A well-ventilated area or a designated collection booth with biosafety features is ideal to reduce transmission risk.

Diagnostic Value

Both NPS and OPS provide suitable diagnostic specimens. Combining both swabs in a single transport tube increases sensitivity, particularly in cases of low viral load.

Materials and Reagents (PPE)

N95 respirator or equivalent particulate respirator

Materials and Reagents (PPE)

Disposable gown or coverall with long sleeves and cuffed wrists

Materials and Reagents (PPE)

Protective goggles or face shield

Materials and Reagents (PPE)

Disposable gloves (*double gloving* recommended in high-risk collection)

Materials and Reagents (PPE)

Head cap and shoe covers (as required by institutional biosafety protocols)

Materials and Reagents (Swabs)

Sterile synthetic fiber swabs (nylon, rayon, or Dacron)

Materials and Reagents (Swabs)

Flexible plastic or wire shafts to prevent trauma during insertion

Materials and Reagents (Swabs)

Not acceptable: *Calcium alginate* swabs and *wooden-shaft* swabs due to PCR inhibition and viral inactivation

Materials and Reagents (Transport Tubes)

Transport Tubes with Viral Transport Medium (VTM) or Universal Transport Medium (UTM)

Components of standard VTM

Hank's Balanced Salt Solution (HBSS) or Phosphate-Buffered Saline (PBS) as the buffer base

Components of standard VTM

Protein stabilizers (0.5-2% bovine serum albumin or fetal bovine serum)

Components of standard VTM

Antimicrobial agents (gentamicin, vancomycin, amphotericin B) to prevent bacterial and fungal overgrowth

Components of standard VTM

Phenol red as a pH indicator

2-3 mL

Tubes should contain __ of VTM/UTM per swab

Labeling and Documentation Materials

Waterproof permanent markers

Labeling and Documentation Materials

Pre-printed or barcoded specimen labels with at least two identifiers (e.g., patient name, date of birth, unique ID)

Labeling and Documentation Materials

Case Investigation Form (CIF) or equivalent requisition form

Ancillary Equipment and Supplies

Sterile tongue depressor (for OPS)

Ancillary Equipment and Supplies

Parafilm or sealing tape to secure transport tube caps

Ancillary Equipment and Supplies

Biohazard specimen transport bags with outer pouches for documentation

2-8°C

Ancillary Equipment and Supplies

Cold chain transport system: insulated box with 4-6 frozen ice packs for ___ storage during transport

Dry ice

Ancillary Equipment and Supplies

___for long-term transport if >3 days (maintain at -70°C)

10% sodium hypochlorite solution and 70% ethanol

Ancillary Equipment and Supplies

Disinfectants: freshly prepared ___ for surface decontamination

Safety Considerations (Infection Risk to Collector)

NPS/OPS procedures bring the collector into close contact with mucosal surfaces and secretions. Aerosols and droplets can be generated by coughing, sneezing, or gag reflex.

Safety Considerations (Infection Risk to Collector)

All specimens must be treated as potentially infectious regardless of clinical suspicion.

Safety Considerations (Required PPE)

N95 respirator mask or equivalent particulate respirator

Safety Considerations (Required PPE)

Eye protection: goggles or full face shield

Safety Considerations (Required PPE)

Disposable fluid-resistant gown with long sleeves and cuffed wrists

Safety Considerations (Required PPE)

Double gloves recommended for higher biosafety during collection

Safety Considerations (Required PPE)

Head cover and shoe covers if required by facility biosafety policy

Safety Considerations (Environmental Controls)

Collection should ideally be performed in a well-ventilated room or dedicated collection booth with negative pressure if available.

Safety Considerations (Environmental Controls)

Limit the presence of nonessential personnel in the collection area.

Safety Considerations (Environmental Controls)

Surfaces should be non-porous and decontaminated with *10%* freshly prepared bleach followed by *70%* ethanol after each collection session.

Safety Considerations (Positioning and Ergonomics)

Collector should stand slightly to the side of the patient to reduce exposure to expelled droplets.

Safety Considerations (Positioning and Ergonomics)

Patients should be seated upright with head tilted slightly back (~*70 degrees*) for NPS or with mouth wide open for OPS.

2-3 mL

Safety Considerations (Specimen Handling and Transport)

Immediately place swabs in sterile tubes containing __ of VTM/UTM.

Safety Considerations (Specimen Handling and Transport)

Ensure caps are tightly closed and secured with Parafilm or sealing tape to prevent leakage.

2-8 C

Safety Considerations (Specimen Handling and Transport)

Place tubes inside biohazard specimen transport bags to the lab ___within 24hours

-70°C

Safety Considerations (Specimen Handling and Transport)

For longer storage (>3 days), specimens must be frozen at __ or transported on dry ice.

Safety Considerations (Post-Collection Precautions)

PPE should be removed in the correct doffing sequence to avoid contamination (outer gloves -> gown -> face protection -> respirator -> inner gloves).

Safety Considerations (Post-Collection Precautions)

Hand hygiene with soap and water or 70% alcohol-based hand rub must be performed immediately after removing gloves.

Safety Considerations (Post-Collection Precautions)

Any accidental spills should be treated with bleach (*10%*) followed by ethanol (*70%*).

Blood

__ is one of the most frequently used specimens in molecular biology because it contains multiple analytes that can be studied at the genomic, transcriptomic, and proteomic levels.

Genomic DNA (gDNA)

Found in nucleated white blood cells (WBCs).

Genomic DNA (gDNA)

Used for genotyping, inherited trait studies, and detection of pathogens integrated in the host genome.

Messenger RNA (mRNA) and Other RNAs

Present in leukocytes but highly unstable due to ribonucleases.

Messenger RNA (mRNA) and Other RNAs

Reflect the gene expression profile of immune and blood cells.

Messenger RNA (mRNA) and Other RNAs

Used for transcriptomic studies, RNA virus detection (HIV, Dengue, SARS-CoV-2), and immune monitoring.

Circulating Cell-Free DNA (ccfDNA)

Composed of short DNA fragments released from apoptotic or necrotic cells.

Circulating Cell-Free DNA (ccfDNA)

Derived from tumor cells, fetal cells, or damaged tissue.

Circulating Cell-Free DNA (ccfDNA)

Valuable for non-invasive prenatal testing (NIPT) and liquid biopsy in oncology.

Plasma Proteins

Plasma contains enzymes, cytokines, antibodies, and other proteins.

Plasma Proteins

Used as biomarkers in immunology and molecular assays like ELISA.

Plasma Proteins

Require stabilization to avoid proteolytic degradation.

Whole blood

__preserves WBCs, the main source of nucleic acids.

Serum

___ formed after clotting, loses leukocytes and yields less DNA/RNA.

Plasma

is superior to serum for viral detection because clots can trap viral particles.

Red blood cells

___ are enucleated and therefore not a source of DNA.

Buffy Coat

The ___ is the thin whitish layer that forms between plasma and red blood cells after centrifugation of anticoagulated blood. It is enriched in leukocytes and platelets, making it a concentrated source of nucleated cells.

DNA and RNA

Because nucleic acids are found within the nuclei of white blood cells, the buffy coat provides higher yields of __ than whole blood. This makes it valuable for molecular applications such as polymerase chain reaction (PCR), sequencing, and transcriptional profiling.

Ficoll-Paque

Isolation of peripheral blood mononuclear cells (PBMCs) is commonly achieved by density gradient centrifugation using ___, a high-molecular-weight polysaccharide.

PBMCs (lymphocytes and monocytes)

When whole blood is layered on Ficoll and centrifuged, ___ form a distinct layer between plasma and Ficoll.

immunological and molecular studies

These PBMCs are crucial for ___, including flow cytometry and gene expression analysis.

variability and biohazard exposure

However, the manual Ficoll method is labor-intensive, requiring careful pipetting and multiple steps that increase __

Blood

provides multiple fractions that are relevant in molecular biology.

White blood cells

White blood cells are the primary source of genomic DNA and RNA.

Plasma

is the preferred specimen for viral nucleic acids and circulating cell-free DNA.

Plasma proteins

serve as important biomarkers for immunologic and molecular assays.

Serum

is less useful for nucleic acid isolation.