Pretransfusion Testing

Pretransfusion testing is performed to select compatible blood components for transfusion, thereby preventing harm to recipient and maximizing posttransfusion survival.

AABB Standards require the following:

1.
Positive patient identification and their corresponding blood specimen;
2.
ABO group and D typing of patient’s specimen;
3.
Testing of patient’s serum/plasma for unexpected, clinically significant RBC antibodies, which are defined as non-ABO antibodies known to cause hemolytic transfusion reactions and/or hemolytic disease of the fetus and newborn (HDFN);
4.
Comparison of current findings with previous results;
5.
Confirmation of ABO group of RBC components (i.e., whole blood and RBCs);
6.
Confirmation of D-type of D-negative RBC components;
7.
Selection of ABO- and D-appropriate components for patient;
8.
Serologic or computer crossmatching (of components containing >2 mL of RBCs); and
9.
Labeling of component for issue, with patient’s identifying information.

Specific tests performed and frequency and methodology of such testing differ based on patient and component to be issued and are done in accordance with the establishment’s standard operating procedures (SOPs).

Patient Identification

Accurate patient identification at time of specimen collection for type and screen and before blood component administration is essential for safe transfusion practice. Mistransfusion is defined as transfusion of blood component (1) with incorrect blood group, (2) intended for another patient, and (3) that was not prescribed. Causes include wrong blood in tube (mislabeling specimen), patient misidentification, and testing errors.

Pretransfusion Orders

Requests must contain two independent patient identifiers to uniquely identify each patient. Other required information on request includes component requested, amount, any special requirements (e.g., irradiation), ordering physician name, and patient age and gender. Patient diagnosis and transfusion and pregnancy history are helpful in guiding the testing and/or component selection.

Type and Screen Request

Type and screen order laboratory to determine the patient’s ABO group, RhD-type, and alloantibody screen. If antibody screen is positive, alloantibody identification is then performed. Patient sample is stored in transfusion service laboratory for crossmatching if transfusion is needed.

Request for Transfusion

If RBCs are requested, appropriate units will be selected and crossmatched. If other components are requested, appropriate units are selected and released.

Patient Specimen

If patient specimen has not been submitted to the laboratory before transfusion request, it must be submitted with transfusion request.

Pretransfusion testing can be performed on either serum or plasma.

Positive patient identification must occur before specimen collection and labeling. If possible, patient should be involved in identity verification process. Specimen must be labeled after collection but before leaving patient’s bedside, with two independent patient identifiers and collection date. In addition, there must be a mechanism to identify phlebotomist (e.g., initial on the specimen label). When pretransfusion specimen is received in the laboratory, an appropriately trained individual must confirm that all identifying information on request is identical to information on specimen itself. In case of any discrepancy or doubt, the specimen is rejected and new specimen must be obtained for testing. Transfusion service should accept only those specimens that are completely, accurately, and legibly labeled with indelible ink. As hemolyzed or lipemic specimens may create problems in evaluating test results, it is important to draw a new specimen whenever possible. Each laboratory must have procedures and policies that define acceptable criteria for accepting specimens for pretransfusion testing and describe how to document and handle unsatisfactory specimens.

Specimen Storage and Retention

Patient specimens and samples of donor RBC components must be stored at refrigerated temperature for at least 7 days after transfusion. This allows for repeat or additional testing if the patient develops transfusion reaction.

Specimen Age

Crossmatching is required for RBC components and other components containing >2 mL of RBCs (i.e., granulocytes). If patient has been transfused or pregnant within past 3 months or transfusion and pregnancy history is unknown, then specimen used for crossmatch must have been collected within 3 days from the time of transfusion event. Each institution should have policy that defines the length of time specimens may be used for patients not transfused or pregnant within the last 3 months. Plasma, platelet, and cryoprecipitate components do not require crossmatching, and therefore, new specimens are not required for these components if results of the patient’s ABO and D typing are on record.

Serologic Testing

In pretransfusion serologic tests, in vitro RBC antigen–antibody reactions are demonstrated, most often, with final endpoint of agglutination. Antibodies against RBC antigens are either IgM or IgG. IgM antibodies can cause direct agglutination of RBCs having corresponding antigens. IgG antibodies usually do not produce direct agglutination after attaching to their corresponding antigens on RBC surface. Antiglobulin test is used to detect RBCs coated with IgG antibodies. This test uses secondary antibody directed against human globulins (anti–human globulin [AHG; also known as antiglobulin]) that attaches and agglutinates sensitized RBCs. AHG can be anti-IgG only or polyspecific containing anti-IgG and anti-C3d and may contain anti-C3b and other Ig and complement antibodies ( Fig. 21.1 ). AHG may be used in direct antiglobulin test (DAT; also known as direct Coombs test) to detect in vivo coating of RBCs or in indirect antiglobulin test (IAT; also known as indirect Coombs test) to demonstrate in vitro reactions between RBCs and antibodies. IAT is used in antibody detection (antibody screen), antibody identification, crossmatching, and RBC phenotyping. Negative AHG test (both DAT and IAT) must be followed by control system of IgG-sensitized RBCs, termed “check cells” (also known as Coombs control), to confirm that the result is not a false-negative. If check cells do not agglutinate, then test must be repeated.

Figure 21.1, Anti–human globulin (AHG) test.

Some RBC antibodies (e.g., anti-Jk ^a and anti-Jk ^b ) are capable of in vitro hemolysis of corresponding antigen-positive RBCs; therefore, reactions should also be evaluated for presence of hemolysis.

Method

Immediate Spin Testing

Either reagent RBCs with known RBC antigen expression are mixed with patient plasma or reagent antisera with known antibody specificity is mixed with patient RBCs. After mixing at room temperature, sample is centrifuged and agglutination or hemolysis is detected.

Indirect Antiglobulin Testing

For IAT, RBCs and plasma or antisera are mixed as mentioned earlier but then incubated at 37°C to enhance binding of IgG. Sample is then washed to remove unbound antibody followed by addition of AHG, centrifugation, and detection of agglutination or hemolysis.

Reactions are interpreted based on degree of agglutination or presence of hemolysis. Alternative methods that do not require agglutination have also been developed, including solid phase and flow cytometry. Many of these are automated.

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