JMIH 2018: How Can We Measure Immune Function in Anoles?

Measuring the swelling induced in anole feet during the PHA assay may result in swelling in one’s own fingers.

The immune system is critical to the survival of animals, including anoles, which are faced with an environment full of potential pathogens and toxins. Ecoimmunologists have developed a myriad of assays to measure various aspects of the immune system and its function in a variety of species, but these assays are often applied to organisms without fully validating them. This issue can prevent a full and accurate interpretation of the results obtained. The PHA skin test is widely used in lizards, including anoles, to test immune function, but has exactly this problem: it has only been validated in cane toads and a crocodile…a large oversight!

Caty Tylan, a PhD candidate and DVM at Penn State University in Tracy Langkilde’s lab, set about rectifying this situation by validating the PHA assay in our favorite squamate lab “rat,” the green anole, Anolis carolinensis. To conduct the PHA test, Caty injected two different types of phytohemagglutinin (PHA-L and PHA-P) into the footpads of green anoles and compared the swelling produced to that of control injections. She also measured types of white blood cells in the blood and foot tissue at regular intervals after injections. Caty found that both types of PHA work well and induce similar levels of swelling with a standard assay protocol in green anoles, but that they induce different types of immune responses. PHA-P elicits a broader response with different types of immune function that varies with time after injection, meaning that the outcomes of this test may be harder to interpret. PHA-L on the other hand, induces higher concentrations of T-lymphocytes, a specific type of white blood cell. As a result, using PHA-L for PHA assays may lead to a test that is more interpretable, especially in studies looking at how the stress response affects immune function.