Serosurveys using Residual Specimens from a Biorepository or Health Facility


This module describes sampling residual blood specimens from biorepositories or health facilities to be used for serosurveys.

Learning Objectives:

  • Define residual blood specimens and identify potential benefits of this design for serological surveillance
  • Describe the ethical, data, and operational considerations prior to accessing a biorepository or health facility specimens
  • Describe considerations for data analysis when using residual blood specimens, particularly their representativeness of the target population

Topics covered in this module
Technical and analytic considerations for accessing and using blood specimens from a biorepository for serosurveys.
Note: Refer to these modules for related information


  • Assay protocols and laboratory data management
  • Using existing clinical specimens from facility or laboratory

Residual or leftover blood specimens collected for other purposes that could be used for serosurveys include:

  • Residual blood specimens available in biorepositories from other surveys, research studies, or surveillance systems
  • Residual blood prospectively collected for clinical purposes at a health facility or laboratory

Although serosurveys using residual blood specimens may be easier and less expensive than community-based serosurveys, there are important considerations and limitations to the use of these convenient specimens (see Table 1 and Figure 1 for more details).



Blood specimens
collected for another purpose

New specimen


Health facility
or laboratory



Interaction with human participants




Design and planning

Moderate: Develop sampling frame and
obtain specimens

Moderate: Identify facilities and
systems to collect specimens and linked data

Moderate: Restricted to specimen-collection

High: All aspects of design and planning required

Data and specimen collection

None (retrospective)

Low (prospective)




Low cost: Specimens collected for other purposes

Moderate: Added costs for specimen collection

High: All general survey and specimen-related costs

Control over Study Design

Little control: Little to no control over data and specimen collection, data availability, data and survey quality


Missing data or specimens could result in bias

Moderate: Less control over the survey design than stand-alone serosurvey, which may result in issues with data availability and quality.

High: Allows for control of study design, sampling,
data and specimen collection, and representativeness of the population of interest



Select existing specimen sources that are representative of the target population,
if possible


Ensure ethical approval allows for further testing of specimens


Consider if seroprevalence may have changed since the specimens were collected.

Additional time, cost, and capacity required to collect specimens (e.g., resources, personnel, community engagement)


Results may not be available in a timely manner



a. Refer to Specimen Collection module for more details.
b. For cost, only the primary drivers are listed.

Table 1. Comparing study designs for serosurveys

When initiating a serosurvey using residual specimens, it is critical to establish a relationship with the individuals or organization responsible for the specimens.  This may be the team who collected the specimens for a research study or surveillance system, or the clinical unit collecting the specimens for another purpose.  Ethical considerations and representativeness of specimens are generally similar for both types of residual specimen sources, whereas specimen considerations are unique to the type of specimen used (Figure 1; see separate sections for more details).

Ethical Considerations
  • Regulatory permissions needed to access residual specimens
  • Approvals for future testing of specimens



Representatives and Potential Biases
  • Target population of interest vs. source of residual specimens
  • Potential biases such that seroprevalence estimates are not representative of target population
  • Individual-level data available for specimens


Specimen Considerations
  • Quality and quantity of the residual specimens
  • Feasibility of accessing residual specimens



Figure 1. Considerations for using residual specimens

Ethical Considerations

  • Additional ethical and regulatory approvals may be required to use specimens collected for another purpose.
  • To determine the feasibility of collecting residual specimens for serosurveys and what ethical approvals are needed, consult with:
    • Health facility administration
    • Lead institution, organization, or individuals that oversee the biorepository
    • Ministry of Health
    • Ethical review boards
  • For biorepositories, refer to the study’s consent form to determine what was agreed upon at the time of providing the specimen. This may include details related to additional testing of the specimens beyond the original purposes and whether there are any limitations to the types of testing that can be performed (e.g., HIV testing).
  • Determine if the results from the serosurvey may be clinically important enough (e.g., disease individual was unaware of) to warrant providing information back to the individual.
  • Individual privacy will need to be respected when accessing patient clinical records for health facility specimens or accessing research study data. Where possible, limit collection of patient identifiers to only those required for the serosurvey research question.  For date of birth, consider converting to age then removing dates.  If any patient identifiers are collected, they must be maintained securely, such as in a locked room or password-protected and encrypted electronic device.

Representativeness and Potential Biases

One of the most important considerations in using residual specimens is the representativeness of the specimen source population in relation to the research question and target population of interest.  It is unlikely that a residual specimen source directly matches the target population so biases will need to be carefully considered and assessments made on what actions could be taken to minimize biases.

Figure 2. Considerations for Representativeness

Refer to Toolkit: Assessing Representativeness for examples highlighting the types of information useful for assessing representativeness and how the information may be used.

Overview of Biorepository Sampling

Selecting a biorepository
To identify an appropriate biorepository for the research question (see Module: Study Design), understanding the source of the specimens is needed to assess the study design. Using the available data linked to the specimens can help assess the representativeness of the biorepository for the target population.

Considerations when selecting a biorepository:
  • How were the specimens originally collected?
    • Examples of biorepository sources:
      • Specimens from previously conducted studies or surveys (e.g., Demographic and Health Survey or Malaria Indicator Survey)
      • Specimens from surveillance systems (e.g., febrile-rash; HIV sentinel surveillance)
      • Specimens originally collected in facilities (e.g., Australian National Serosurveillance Program [ANSP])
    • It is important to ensure the age groups and populations align with the population of interest.
    • If testing requires a particular specimen type (e.g., serum or plasma), this could limit what biorepositories are available.
  • What were the eligibility criteria for the biorepository?
    • If inclusion/exclusion criteria were narrow in development of biorepository, this could limit generalizability.
      • E.g. Research question is for childhood diseases but specimens were only collected from adults
      • E.g. Research question is for disease that affects males, but specimens were only collected from antenatal care clinic
  • How long has it been since the specimens were collected?
    • If there has been an epidemiological change, such as vaccine introduction or large-scale outbreaks, this could affect seroprevalence estimates. This may include large-scale impacts on the health system, such as disruptions related to SARS-CoV-2.
  • How much of the biorepository will be needed?
    • If a large biorepository is available, consider what age groups, geographic locations or other demographic characteristics could guide sampling.

Because specimens were previously collected, you will need to evaluate the conditions under which they have been maintained. Specimens should be clearly labeled with a unique identifier, organized in a meaningful way to facilitate identification, and listed in a database that contains freezer location information as well as links to other demographic data.
The conditions required for specimen storage will vary depending on the specimen type. For specimens that have been frozen, monitoring freezer temperatures, the number of times the specimen aliquots have been thawed and refrozen, and how long they have been stored are important in assessing specimen quality and whether serological testing is likely to yield accurate results.
The completeness of the biorepository should be assessed. Some specimens may be missing, misplaced or have insufficient sample volume remaining. If a high percentage of specimens are missing or missing specimens share characteristics (i.e., missingness is not random), this could lead to biased seroprevalence estimates. Conducting a missingness analysis can help assess whether there may be biases in the sampled specimens.

Once an appropriate biorepository has been identified and specimens can be accessed, the following steps can be taken:

Figure 3. Steps in conducting a serosurvey using specimens from a biorepository

Once specimen testing is complete, analysis of data would proceed as for any serological survey. Analysis considerations will vary based on study design, such as whether the specimens were collected using probability-based sampling. If probability-based sampling was done, consider analytic techniques such as weighting, propensity scores, or multiple imputation methodologies. Some aspects to consider to enhance generalizability to the target population:

  • Probability of selection—Consider both the chance that a specimen was included from the original study population and the chance that a specimen from the biorepository was chosen for the serosurvey.
  • Non-response—If missing specimens are a non-random sample of the biorepository, consider weighting to align the sample with the overall biorepository.
  • Post-stratification weighting—Can be done to ensure the tested specimens match the characteristics of the target population.

For non-probability sampling, consider what adjustments could be made to the sample population to be more representative of the target population (e.g., adjust age distribution of the sample to reflect that of the target population).
Other analytical techniques could be appropriate for your study design. Consultation with a biostatistician or further reading can help. Refer to the WHO Vaccination Coverage Cluster Survey manual for more detailed information about sample sizes (Section 2.7) and analyses (Section 6).

Overview of Sampling Residual Specimens from Facilities

Selecting health facilities
To select potential health facilities for prospective residual blood specimen collection, it is important to gather background information to assess the feasibility of collecting residual blood specimens and inform serosurvey procedures. This includes gathering information on the type and location of health facilities, types of patients using the facility, availability of residual specimens, types of laboratory tests performed, procedures for collecting, storing, and testing specimens, and demographic data available for each specimen. Detailed considerations are provided in box below. Refer to Toolkit: Facility Characteristics Form which is an example of a detailed form to collect background information on health facilities for residual blood specimen collection.

Considerations when selecting facilities:
  • Is the facility public or private? Is it a medical facility or a diagnostic laboratory?
  • Who are the patients using the facility (urgent care, primary care, tertiary care; specialized care)?
  • Are the patients representative of the population of interest (e.g., similar age and geographic distribution)? What types of patients typically have blood drawn at these facilities (inpatient, outpatient, chronic vs. acute conditions, emergency, trauma, etc.)?
  • Is there the potential for specimens to be received from patients living outside the study area? If yes, can those specimens be identified?
  • What types of tests are typically performed on the specimens (routine blood panel, biochemistry, specialized tests)? What types of specimens are collected for each type of test (whole blood, serum, or plasma)? Are all specimens tested onsite or are some specimens sent to an external lab for testing?
  • For large hospitals, are there multiple laboratories, and if yes, which departments are served by each laboratory?
  • How long does the facility retain the specimens? What are the storage conditions while the facility is holding the specimens? What is the maximum time the specimens that could be stored?
  • How are the specimens linked to the data (by name, by hospital ID, etc.)? How is the data linked to specimens recorded (paper-based or electronic or both)? Is the information available in a single database or separate sources?

There are two primary survey staff that can be involved in collection and transportation of residual specimens:

Site Coordinator
  • Coordinates all staff at the site and communications with all facilities providing specimens.
Collection Staff
  • Identifies and collects specimens at facilities.
  • Abstracts data about each specimen from facility register(s) and enters data into an electronic database. ​
  • Transports specimens to the laboratory where they will be processed and stored until testing.

Figure 4. Roles and Responsibilities for Sampling from Health Facilities

Ideally survey staff are hired to coordinate the project and work closely with the health facility staff. Depending on the facility’s procedures and capacity, the specimen collection and data abstraction may be done by the facility staff (following training by the survey team) or by survey staff themselves.

Specimen collection procedures should be tailored to the facility so collection can be integrated within standard procedures. Below is a general overview of the steps in residual specimen collection.

a. Refer to the blood specimen processing, storage, and shipment and assay protocols and laboratory data management modules for more information.

Figure 5. Steps in conducting a serosurvey using residual blood specimens. Steps in orange are conducted by the health facility laboratory and steps in blue are completed as a part of the serosurvey.

Protocols will differ depending on specimen type, sampling method used and the facility-specific procedures. Refer to Toolkit: Facility Serosurvey Specimen SOP for a detailed protocol and supply list for collecting sera from a health facility using residual blood specimens.

Only residual specimens meeting selection criteria established by the serosurvey should be collected and tested. Example selection criteria include specified age range, specimen type (whole blood, serum, or plasma), minimum volume (as determined by the volume required for the assay), or absence of hemolysis. See below for additional considerations for collecting residual specimens.

Considerations for collection of residual specimens:
  • Time-frame for eligible specimens (e.g., within a prespecified number of days of collection) will depend on the facility’s procedures for retaining specimens (including storage conditions) and the specimen type.
  • Sampling strategies may be used to reduce bias and account for availability of specimens, including:
    • If the number of residual specimens is limited at a facility, collection staff may collect all specimens acknowledging that some may have insufficient volume after processing.
    • If the number of residual specimens is large at a facility, collection staff may use quota sampling, e.g., only collecting the first X specimens during each visit, where X depends on sample size, how often specimens are collected, and how many specimens can be feasibly collected in a visit.

Figure 6. Identifying eligible residual specimens from health facility

After specimens are collected from the health facility, the specimens should be transported to the laboratory for further processing, storage and testing. Refer to Facility Serosurvey Specimen SOP, and Specimen Processing and Assay protocols modules for detailed information on specimen processing and testing.

Data requirements may vary by survey objectives but the key variables to capture related to specimen characteristics and processing include:

Specimen Characteristics
  • Study ID (assigned by survey staff)
  • Facility location
  • Originating facility (if specimens provided by a diagnostic laboratory)
  • Age
  • Sex
  • Collection date
Specimen Processing
  • Study ID (assigned by survey staff)
  • Processing date
  • Volume (uL)
  • Hemolysis (e.g., none, mild, moderate or severe)
  • Location of specimen (e.g., aliquot freezer box and box position)

Figure 7. Key variables for residual specimen collection.

The data may be captured in a paper-based register while at the health facility or laboratory and then entered into an electronic database.

Depending on the survey objectives, additional data may be valuable, including reason for specimen collection and information on village, town or district of residence to evaluate the geographic distribution of specimens and to determine whether the specimens are representative of the target population.  However, these data points may not be available at the laboratory, requiring additional steps by to gather this information.

Considerations when developing facility-specific protocols include the movement of specimens from the point of collection to testing, availability of data to identify eligible specimens, and type of data records at hospital (e.g., paper vs electronic, single or multiple databases). Refer to Toolkit: Residual Specimen Case Studies and Toolkit: Facility Serosurvey Specimen SOP for detailed facility specific protocols.

Examples of Serosurveys Using Residual Specimens

Example of Sampling from Biorepository


Section Toolkit material
Representativeness and Potential Biases Toolkit Assessing Representativeness
Sampling Residual Specimens from Facilities Facility Characteristics Form
Sampling Residual Specimens from Facilities Facility Serosurvey Specimen SOP
Sampling Residual Specimens from Facilities Facility Collection Case Studies