Module 6 · Chapter 7
Specimen Collection and Handling for AEFI
Biological samples, the national virology labs, CDL Kasauli for vaccines and diluents, CDL Kolkata for syringes and vitamin A — quantities, sealing, packing and shipping.
Overview
Samples that may be tested during an AEFI investigation:
- Biological samples from the patient, and
- Vaccines, diluents and logistics such as syringes (only where indicated).
Understand the need clearly before collecting any specimen. Specimens must travel with supporting documents — Lab Request Form (LRF), CRF, CIF and other records — stating the tests required and any specific requests.
7.1 Testing of biological specimens
- Usually decided by the treating physician based on signs, symptoms and differential diagnosis (which sample, which test, where).
- The District AEFI Committee/DIO may recommend tests in identified government or accredited private labs (approved by the competent authority).
- National labs can tell vaccine virus from wild virus:
• AEFI after JE vaccine → CSF and blood to NIV Pune or Gorakhpur
• AEFI after MR vaccine → NIV Mumbai - Label, pack and transport properly with LRF, CRF, CIF.
Table 7.1 · National laboratories
| Laboratory | Address | Contact |
|---|---|---|
| ICMR-National Institute of Virology (JE Group), Pune | The Director, 20/A Dr Ambedkar Road, Pune, Maharashtra 411001 | director.niv@icmr.gov.in · 020-26006290 / 25906890 |
| ICMR-RMRC Gorakhpur (formerly ICMR-NIV Gorakhpur Unit) | BRD Medical College campus, Gorakhpur, UP 273013 | admn-rmrcgkp@nic.in, admnrmrcgkp@gmail.com · 0551-2971007 |
| ICMR-NIV Mumbai Unit (formerly Enterovirus Research Centre) | Haffkine Institute Compound, A.D. Marg, Parel, Mumbai 400012 | nivmumbaiunit@gmail.com · 022-24134130 / 24125309 |
Table 7.2 · Responsibilities for biological specimens
| Activity | Responsibility |
|---|---|
| Decision to collect (as soon as possible) | Treating physician; DAC/DIO may recommend certain tests |
| Laboratory | Identified government and accredited private labs (approved by competent authority) |
| Funding | AEFI surveillance, case management and related activities: PIP FMR code RCH 4 – Immunization – S. No. 32 – SSRE (Surveillance Research Review Evaluation). NIV Pune, Gorakhpur and Mumbai bear testing costs for AEFIs after JE and MR vaccines. |
| Reporting results | Labs send results to everyone whose contact details (full address with PIN, phone, fax, email) are in the LRF |
Table 7.3 · Biological specimens and tests
| Event | Specimen | Tests |
|---|---|---|
| Severe local reaction · abscess · lymphadenitis | Blood | Culture |
| Pus | FNAC (as listed) | |
| CNS symptoms (with/without paralysis) | Serum | IgM and IgG antibodies for viral pathogens |
| Blood | Culture | |
| Cerebrospinal fluid | Differential cell count, biochemistry, bacterial and viral culture, PCR (HSV-1/2, enterovirus, other) | |
| Stool* | Enterovirus and viral culture | |
| Anaphylaxis | Serum | Mast cell tryptase; specific IgE |
| AEFI after COVID-19 vaccination | Nasal swab | RT-PCR for COVID-19 |
*If paralysis follows OPV, stool specimens are important — collect as per AFP stool-collection guidelines.
7.1.1 Post-mortem specimens
- Perform the post-mortem as soon as possible to avoid tissue damage, artefacts and lysis of the adrenal glands.
- DIO includes a detailed history in the post-mortem form so the team looks for underlying disease.
- Send histopathology and toxicology samples to approved/accredited government reference labs through the investigating police, early, properly labelled, with the post-mortem report listing tests required.
- The report should ideally list cause of death per ICD-10 and, if possible, causative agents/drugs (Annexure 12).
7.2 Testing of vaccine/diluents at CDL Kasauli
What CDL Kasauli testsAll vaccines and diluents are tested for physical aspects, sterility, abnormal toxicity and biochemical identity. Potency is not tested in AEFI cases. Results are sent in about 30 days.
Request testing only on a clear suspicion — cluster of AEFIs, increase in vaccine reactions, public/community concern — never routinely and never before a working hypothesis.
7.2.1 Sample collection
- The DIO and drug inspector collect adequate quantities of the implicated vaccine/diluent from the AEFI site and the last vaccine storage point, and ship in cold chain to CDL Kasauli as early as possible.
- Form 18 is used when drug inspectors send samples; the LRF when the DIO sends them.
- Prepare four sealed sets of equal quantity: 1 → CDL Kasauli; 1 retained at the site of collection (PHC/CHC or district HQ); 2 retained with the drug inspector.
- Collect from the next available storage point if needed; for freeze-dried vaccines collect the same quantity of diluent. The quantity required by CDL must not be compromised.
Table 7.4 · Quantity of implicated vaccine/diluent to collect
A = unused vaccine vials collected · B = unused diluents collected · C = vials shipped to CDL (¼ of A) · D = diluents shipped (¼ of B)
| Vaccine | A · collect vials | B · collect diluents | C · ship vials | D · ship diluents |
|---|---|---|---|---|
| DPT group (incl. pentavalent) | 10-dose × 40 OR 1-dose × 120 | NA | 10-dose × 10 OR 1-dose × 30 | NA |
| BCG | 10-dose × 160 OR 20-dose × 160 | 160 | 10-dose × 40 OR 20-dose × 40 | 40 |
| OPV | 20-dose × 40 | NA | 20-dose × 10 | NA |
| Measles / MMR / MR | 1-dose × 80 OR 5-dose × 60 OR 10-dose × 40 | 80 / 60 / 40 | 1-dose × 20 OR 5-dose × 15 OR 10-dose × 10 | 20 / 15 / 10 |
| JE & hepatitis vaccines | 1-dose × 80 OR 5-dose × 60 OR 10-dose × 40 | 80 / 60 / 40 | 1-dose × 20 OR 5-dose × 15 OR 10-dose × 10 | 20 / 15 / 10 |
| IPV | 5-dose × 40 OR 10-dose × 40 OR 1-dose × 80 | NA | 5-dose × 10 OR 10-dose × 10 OR 1-dose × 20 | NA |
| Rabies | 1-dose × 60 | 60 | 1-dose × 15 | 15 |
| Rotavirus (oral) | 1-dose × 80 OR 2-dose × 80 OR 5-dose × 60 OR 10-dose × 40 | NA | 1-dose × 20 OR 2-dose × 20 OR 5-dose × 15 OR 10-dose × 10 | NA |
| Pneumococcal | 4-dose × 60 | NA | 4-dose × 15 | NA |
| COVID-19 | 10-dose × 8 OR 20-dose × 5 | NA | 10-dose × 8 OR 20-dose × 5 | NA |
Note the exceptionFor COVID-19 vaccine the quantity collected and the quantity shipped are listed as the same (10 × 8 or 20 × 5) — unlike the “¼ of total” rule for other vaccines.
Table 7.5 · Responsibilities — vaccine/diluent/syringe samples
| Activity | Responsibility |
|---|---|
| Decision to collect | District AEFI committee (includes local drug inspector); consult state AEFI committee if required |
| Collecting and sending samples | Drug inspector and DIO |
| Sample types | As recommended by the DAC; the drug inspector may collect additional samples |
| Packaging and cold chain | Drug inspector and DIO |
| Sealing with “official lac seal” | Preferably drug inspector; if unavailable, the CMO’s seal |
| Transport to lab | Preferably DIO and/or drug inspector |
| Laboratory | Identified laboratories in this chapter |
| Funding | Surveillance, case management, transport: PIP FMR RCH 4 – Immunization – S. No. 32 – SSRE. Actual testing at CDL Kasauli and Kolkata is borne by the laboratories. NIVs bear JE/MR testing costs. |
| Reporting results | Lab forwards copy to CDSCO, AC Immunization Division MoHFW, State Immunization Officer, State Cold Chain Officer and State drug authority, and to everyone listed in the LRF |
| Feedback of results | DIO shares with the drug inspector, reporting MO and reporting private facility |
| Temporarily suspending the implicated batch (only on suspicion of quality — very rare) | Local drug authority with DIO and DAC, in consultation with SEPIO/State AEFI Committee and MoHFW |
7.2.2–7.2.3 Packing, documentation and transport
- Separate plastic zipper bags for each vaccine and diluent; label each with name, age, date of collection, AEFI EPID number and point of collection.
- Put all bags (vaccines and diluents separately) into a bigger zipper bag → cardboard box, tied with string on all sides, with an “official lac seal” by the drug inspector (or CMO) — Figure 7.1.
- The completed LRF (Annexure 13) is sealed with the same lac seal (prevents tampering); CRF and CIF accompany the samples.
- Vaccines and diluents are tested together → send diluents of freeze-dried vaccines (BCG, MR, JE) with the vaccine.
- Transport under cold chain (vaccine carrier or thermocol box with ice packs), preferably by messenger.
- CDL Kasauli accepts samples all days of the week; the messenger must insist on a “sample received receipt” (shows sample condition; not issued on weekends).
- Experienced courier services that guarantee delivery time and condition may also be used.
Address: Director, Central Drugs Laboratory, Central Research Institute, Kasauli – 173204, Himachal Pradesh · nclkasauli@gmail.com · 01792-272578 / 272046
Sending opened/used vials
Used (opened) vials are not technically required, but sending one (if available) helps confirm the same batch is tested. Opened vials usually aren’t tested because: quantity often inadequate; they become unsterile once opened; reconstituted vials cannot be tested beyond 4 hours; opened vials have weak legal sanctity.
Under the Open Vial Policy, vials returning from a session are segregated into OVP and non-OVP groups. Non-OVP vials go into a plastic box marked “not to be used” in the ILR and are discarded after 48 hours or before the next session, whichever is earlier — but not discarded if an AEFI is reported.
Worked example (MR, district M)
5-dose MR vial AEFIThe DAC decided to test the implicated MR batch. Per Table 7.4 the DIO and drug inspector needed 60 vials + 60 diluents. At the session site they found only one partial and one unused vial with the ANM, so they collected 59 unused vials from the PHC storage point to complete 60. Diluents: 45 from the PHC + 15 from the district vaccine store = 60. They made four equal sets of 15 vials + 15 diluents (eight zipper bags, labelled by point of collection). One set was sealed with the drug inspector’s lac seal and sent under cold chain to CDL Kasauli with LRF, CRF and CIF; the rest were retained as per guidelines.
7.2.4 Dos and don’ts
Dos
- Collect unused samples only from the implicated batch.
- Send samples with the official lac seal.
- Seal the LRF with the lac seal too.
- Pack diluents carefully and separately.
- Mark the point of collection on each zipper bag.
- Keep manufacturer labels (name, batch, mfg/expiry) intact and visible.
- Pack to avoid breakage (manufacturer cartons); zipper bag → vaccine carrier/thermocol with ice packs. Dry ice may be used for OPV; NEVER for freeze-sensitive vaccines.
- Write CDL Kasauli’s address clearly on the box.
- Send with LRF and CRF; CIF and other records if available.
Don’ts
- Never wrap labels with adhesive tape or cover them with other labels (Figure 7.2).
- No wetting or mutilation of labels — put extra labels on the zipper bags instead.
- Vaccines must not be expired when they reach the lab.
7.3 Testing of syringes, needles and vitamin A at CDL Kolkata
- CDL Kolkata tests implicated AD syringes, reconstitution syringes and vitamin A for standard sterility and physical parameters.
- Only after a decision by the district or state AEFI committee and/or a clear basis of suspicion — not routine. Results in about 60 days.
- The drug inspector collects samples per Drugs and Cosmetics Rules; samples must be of the same manufacturer and batch.
- Four equal sets: 1 sent for testing, 1 at the point of collection, 2 with the drug inspector. Send by reliable courier or post — cold chain NOT required.
Table 7.6 · Quantity of unused samples (implicated batch)
| Sample | Quantity |
|---|---|
| AD syringes | 4 sets × 50 (total 200): 50 to CDL Kolkata; 50 at source; 2 × 50 (100) with drug inspector |
| Reconstitution syringes | 4 sets × 50 (total 200): same distribution |
| Vitamin A | 4 sets × two 100-ml bottles (total 8): 2 bottles to CDL Kolkata; 2 at source; 4 with drug inspector |
7.3.2 Packing, documentation, shipment
Send used samples (if available) along with unused ones of the same manufacturer, sealed in separate labelled packets, in a cardboard box tied with string and sealed with the drug inspector’s (or CMO’s) lac seal. Send with completed LRF and CRF; CIF if requested. A used vitamin A bottle may accompany unused sealed bottles, packed against breakage/spillage.
Address: The Director, Central Drug Laboratory, MoHFW, 3 Kyd Street, Kolkata – 700016 · cdlkol@cdsco.nic.in · Phone 033-22299541 · Fax 033-22299380
Important considerationsHealth authorities must coordinate with the police/other investigating agencies and acquaint them with the national AEFI guidelines. All original documents are retained by the MO in charge; agencies receive attested copies.
Chapter summary
- Biological samples and vaccines/diluents/logistics (syringes, vitamin A) can be tested.
- Biological testing is the treating physician’s decision.
- Vaccine/logistics testing only on clear suspicion — clusters, increased reactions, community concern.
- DIO and drug inspector collect adequate quantities.
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