10.1 Procedural Definition: What Is This Test About?

True immune‐mediated erythrocyte agglutination can be seen in primary and secondary immune‐mediated hemolytic anemia. When aggregation of erythrocytes is seen macroscopically in the blood collection tube or microscopically on examination of the blood film, it is important to determine if the aggregation is antibody mediated or nonspecific adherence. The saline agglutination test is used to determine whether erythrocyte agglutination is due to antibody‐specific cross‐linkage or nonspecific adherence due to high protein or altered proteins in the plasma. The principle of the test is to dilute the blood sample and observe whether or not the erythrocytes disperse into individual cells or if they remain aggregated.

10.2 Procedural Purpose: Why Should I Perform This Test?

Immune‐mediated hemolytic anemia is not uncommon in the dog but less common in the cat. In some forms of immune‐mediated hemolytic anemia, antibody‐dependent agglutination of the erythrocytes is seen. In patients that present with a regenerative anemia and gross or microscopic evidence of erythrocyte agglutination, it is essential to confirm that the agglutination is antibody mediated. High plasma protein or increased concentration of inflammatory proteins with higher affinity for the erythrocyte surface can result in nonspecific erythrocytes aggregation. The saline agglutination test is an inexpensive screening test that can assist in differentiating these two mechanisms of erythrocyte aggregation or agglutination.

Immune‐mediated anemia in the dog and cat can be a primary or secondary disease. Primary immune‐mediated disease is diagnosed by confirming antibody‐mediated removal or destruction of erythrocytes, ruling out secondary disease as a cause of the immune‐mediated erythrocyte destruction and ruling out nonimmune causes of the agglutination.

The saline agglutination test has a long history of use in veterinary medicine. However, it is somewhat controversial. The protocol for the test has not been standardized.

10.3 Equipment

Equipment used for this test include:

• whole blood in blood collection tube with anticoagulant
  
• glass slides
  
• pipette
  
• saline (0.9% NaCl)
  
• coverslips
  
• gloves
  
• Optional: clean conical centrifuge tube.

10.4 Procedural Steps: How Do I Perform This Test?

• Gather all the necessary supplies for the test (see Figure 10.1).