Histamine as an aid to biopsy of third eyelid lymphoid tissue in sheep

ers (1998b, 2000), is a practical test for live sheep and is useful for the preclinical diagnosis of scrapie in surveillance programmes (O'Rourke and others 2002). Although the lymphoid tissue of the third eyelid is accessible and can be anaesthetised locally, one obstacle to using this diagnostic test is obtaining third eyelid samples with sufficient numbers of lymphoid follicles to enable accurate scrapie diagnosis. This is complicated by the irregular distribution of the lymphoid tissue within the third eyelid and the sparse amount of lymphoid tissues in older sheep. A previous study reported lymphoid follicle mean (se) numbers of 0-7 (0-16), 0-78 (0.27) and 5-32 (0.7) in three areas of the third eyelids of sheep (Thuring and others 2000). Suitable samples must contain at least six lymphoid follicles per section for the diagnosis of scrapie (O'Rourke and others 2002). This short communication reports an improved third eyelid sampling technique using histamine/proparacaine eye drops. A field trial was conducted on 78 sheep from a Navajo reservation flock in Alamo, New Mexico, using an eye drop formulation of 1 per cent histamine and 0.5 per cent proparacaine hydrochloride in an aqueous base (Vetricare) before third eyelid biopsy. Seventy-six of the sheep (97.4 per cent) provided sufficient numbers of lymphoid follicles in their third eyelids. No adverse reactions to the histamine or biopsy procedure, such as photophobia, increased attraction of flies, excessive tearing or bleeding from the biopsy site, or anorexia, were noted on the day of testing or during follow-up care conducted on site the next day. The flock was checked on a regular basis for the following 12 months and no adverse reactions from the biopsy protocol were noted. Five additional sheep flocks on the Navajo Nation reservation have been third eyelid tested to date using the histamine/proparacaine eye drops. The only reported side effect has been mild bleeding from the biopsy site in a small number of sheep. In each of these cases, the bleeding stopped within a few minutes. In total, 131 sheep were biopsied using the histamine/ proparacaine drops, of which 13 gave insufficient numbers of follicles for scrapie diagnosis. Data from 53 control sheep, which had been biopsied with proparacaine drops alone with the use of no added histamine, showed 16 biopsy samples with insufficient numbers of follicles for scrapie diagnosis. An analysis of these data show that the success rate for obtainFIG 1: Immunoreactivity for PrPSc (red) detected in lymphoid tissue of third eyelids treated (a) without and (b) with histamine-containing eye drops. x 40