What Factors Are Considered During Serology Testing?

Serology testing is a method used to detect specific molecules within a blood sample, primarily in the liquid component known as serum. Its purpose ranges from diagnosing infections and evaluating immune responses to vaccines, to screening for autoimmune diseases. The reliability of these tests depends on evaluating multiple scientific factors. Accurate interpretation requires understanding these factors to ensure the results are meaningful and correctly applied.

The Specific Target: Antigens and Antibodies

Serology testing is designed to find a specific target, which is either an antigen or an antibody. Antigens are foreign substances, such as proteins on the surface of a virus or bacterium, that provoke an immune response. Detecting these antigens can indicate an active infection. In contrast, antibodies are proteins produced by the immune system to identify and neutralize these invaders. The presence of antibodies reveals an immune response to a pathogen from a past infection or vaccination.

Different pathogens or medical conditions necessitate searching for distinct targets. For instance, tests might look for specific antibodies like Immunoglobulin M (IgM) or Immunoglobulin G (IgG) to help determine the stage of an infection. For autoimmune conditions, the test looks for autoantibodies, which are antibodies that mistakenly target the body’s own tissues. The precision of the test depends on its ability to bind only to the intended target, avoiding cross-reactivity with similar molecules that could lead to inaccurate results.

Timing of the Test: Capturing the Immune Response

The timing of a serology test in relation to a suspected infection or vaccination is a major factor in its accuracy. The immune system needs time to generate detectable antibodies following exposure. Testing during the initial “window period,” when levels are too low to be measured, can produce a false-negative result even if the person is infected.

The production of different antibody classes follows a general timeline. IgM antibodies are the first to appear, often becoming detectable within a week of symptom onset, but their levels usually decline over several weeks. Following the initial IgM response, IgG antibodies begin to appear, generally around seven to 14 days after infection. IgG levels increase and often remain elevated for a much longer duration.

This dynamic allows clinicians to differentiate between infection stages. A positive IgM result with a negative IgG result might suggest a very recent infection, while positive results for both can indicate an ongoing infection. The presence of only IgG antibodies points to a past infection or successful vaccination, as the short-lived IgM antibodies have waned. For example, with rubella, IgG antibodies can last a lifetime, while IgM persists for up to three months.

A single test provides a snapshot, but sometimes paired samples, taken weeks apart, are needed to confirm a recent infection by observing if antibody levels are rising. For some diseases, the optimal time to collect a sample is five days after symptoms begin, as more than 90% of cases will be IgM positive by then. If an earlier sample is negative, a second one may be required to rule out the infection.

Patient-Specific Influences

A patient’s unique characteristics can significantly influence serology test results. A person’s immune status is a primary consideration, as immunocompromised individuals may not produce antibodies at the same rate or level as healthy people. Conditions like hematological malignancies or the use of immunosuppressive medications can diminish the antibody response, potentially leading to lower-than-expected results.

Age also affects immune function. Infants have immature immune systems, and older adults may experience immunosenescence, a gradual decline in immune effectiveness. Both groups can have altered antibody responses to infection or vaccination, which must be considered when interpreting results. The age when recurrent infections begin can offer clues; for instance, onset before six months of age may suggest a T-cell defect.

A patient’s history of prior infections or vaccinations is another factor. Previous exposure to related pathogens can sometimes lead to cross-reactive antibodies that are detected by a test for a different organism, causing a false-positive result. Vaccination history, including the type of vaccine and the time elapsed since it was administered, directly impacts the antibody profile a serology test will measure.

Assay Characteristics and Methodology

The technical specifications of the serological test, known as the assay, are fundamental to interpreting its results. Two primary characteristics define a test’s performance: sensitivity and specificity. Sensitivity is the test’s ability to correctly identify individuals who have the target molecule (true positives). Specificity is the test’s ability to correctly identify individuals who do not have the target (true negatives).

Several different methodologies are used for serology testing, each with its own performance profile.

• Enzyme-linked immunosorbent assays (ELISA) are a common, lab-based method known for high sensitivity and specificity.
• Lateral flow assays (LFA) are often used in rapid, point-of-care tests and provide quick results but can have lower sensitivity.
• Chemiluminescence immunoassays (CLIA) are another widely used automated lab method.
• Neutralization assays measure the functional ability of antibodies to block a pathogen.

The choice of method depends on the clinical context, such as the need for speed versus the demand for high accuracy. For example, one study found an ELISA had 94.0% sensitivity and 98.0% specificity, whereas a lateral flow assay showed 78.0% sensitivity and 90.5% specificity for the same antibodies. These differences highlight why the specific assay used must be considered when evaluating a result. Every assay also uses a predetermined cut-off value to distinguish between a positive and a negative result, and values close to this threshold may require retesting.

Sample Integrity and Pre-Analytical Variables

Pre-analytical variables related to the blood sample can impact test results. The type of sample required—whether whole blood, serum, or plasma—is specified by the test manufacturer, and using the wrong type can lead to inaccurate outcomes.

Proper collection and handling procedures are necessary to maintain sample integrity. Poor blood drawing technique can cause in-vitro hemolysis, the rupture of red blood cells. This releases cell contents into the serum or plasma, which can interfere with the chemical reactions in many assays and lead to erroneous results or canceled tests.

Lipemia, a high concentration of fats in the blood, can also interfere with tests. Often caused by a patient not fasting, it makes serum appear milky and can affect the light-based measurements used in many automated analyzers. While some interference from lipemia can be resolved through special laboratory procedures, it remains a significant pre-analytical variable.

Sample transport and storage temperature, along with the time between collection and testing, are also important. Prolonged delays or improper temperatures can lead to the degradation of target molecules. Adhering to strict protocols for sample handling is a basic requirement for reliable serological testing.