for virus does not indicate whether a viral infection took place recently or not.

The ELISA test is the blood test most commonly used for the diagnosis of toxoplasmosis and toxocariasis. The presence of a high IgM anti-Toxoplasma and anti-Toxocara titers indicates a recent infection.

3.2.1.9 Interferon-Gamma Release Assays (IGRAs)

Recently, two in vitro assays that measure T-cell release of interferon-gamma (IFN-g) in response to stimulation with the highly tuberculosis-spe- ciÞc antigens, ESAT-6 and CFP-10, have become commercially available [74]. QuantiFERON-TB Gold¨ (Cellestis Ltd., Carnegie, Australia) is a whole-blood ELISA for measurement of IFN-g, and T-SPOT.TB¨ (Oxford Immunotec, Oxford, UK) is an enzyme-linked immunospot (ELISpot) assay. IGRAs are more speciÞc than the intracutaneous tuberculin puriÞed protein derivative (PPD) as a result of less cross-reactivity due to BCG vaccination and sensitization by nontuberculous mycobacteria [75]. IGRAs also appear to be at least as sensitive as the PPD for active tuberculosis (used as a surrogate for latent

Mycobacterium tuberculosis infection). Although diagnostic sensitivity for latent infection cannot be directly estimated because of the absence of a gold standard, these tests have shown better correlation than the PPD with exposure to M. tuberculosis in contact investigations in low-incidence settings.

Other potential advantages of IGRAs include logistical convenience, need for fewer patient visits to complete testing, avoidance of unreliable and somewhat subjective measurements such as skin induration, and the ability to perform serial testing without inducing the boosting phenomenon (a spurious PPD conversion due to boosting of reactivity on subsequent PPDs among BCGvaccinated persons and those infected with other mycobacteria). Because of the high speciÞcity and other potential advantages, IGRAs are likely to replace the PPD for latent infection diagnosis in low-incidence, high-income settings, where cross-reactivity due to BCG might adversely

impact the interpretation and utility of the PPD. Direct comparative studies in routine practice thus far suggest that the ELISpot has a lower rate of indeterminate results and probably a higher degree of diagnostic sensitivity than the wholeblood ELISA. Further studies are under way to assess the performance of these tests in contact investigations and in persons with suspected tuberculosis disease, health care workers, HIVinfected individuals, persons with iatrogenic immunosuppression, and children. These tests may be useful for antituberculous treatment decision making in chronic ocular diseases [76Ð78].

3.2.2Anterior Chamber Polymerase Chain Reaction Testing

Polymerase chain reaction (PCR) of aqueous samples obtained by anterior chamber tap (approximately 100Ð200 ml volume) can be useful in patients with sclerouveitis for detection of DNA from herpes simplex virus, VZV, cytomegalovirus, and for the parasite Toxoplasma gondii. This procedure can be performed conveniently in the outpatient setting and has been shown to be safe in the hands of an experienced ophthalmologist dealing with uveitis [79].

3.2.3Smears and Cultures

Scrapings for smears and cultures must be obtained in cases of infectious suspect. Material from vigorous scraping of the infected scleral or corneoscleral area with a surgical blade should be smeared onto glass slides for staining (Gram and Giemsa) and onto agar plates or broths for bacterial or fungal cultures (two blood agar preparations: one kept at 35¡C for blood agar, chocolate agar, Sabouraud dextrose agar, thioglycollate broth, and brainÐheart infusion medium and the other kept at room temperature). In case of acanthamoeba suspect, staining with calcoßuor white stain or Gomori methenamine silver and culture in non-nutrient agar with Escherichia coli must be performed.

forms of episcleritis and scleritis, providing considerable information in guiding subsequent therapy [81, 82].

In our experience, corneal involvement can be found in 13% of patients with scleritis. Although most of the different forms of keratitis associated with scleral disease can be diagnosed clinically, differentiation between the early changes of the relatively benign peripheral corneal opaciÞcation with or without neovascularization and the early changes of the more serious corneal thinning, either with limbal guttering or peripheral corneal ulceration, can sometimes be difÞcult. Early detection of the most severe forms of keratitis associated with scleritis is important if one is to institute adequate treatment before visual acuity becomes affected. Because the accurate diagnosis of keratitis associated with scleritis can add valuable information to the choice of therapy, it is important to Þnd objective methods to evaluate early keratitis. Anterior segment ßuorescein angiography can sometimes help in this regard [83].

Adequate medical treatment for scleral inßammation with or without corneal involvement frequently results in halting the process, either through new vessel formation to cover the defect or through recanalization of existing vessels [84]. Although most of the individual responses to treatment can be easily monitored by clinical examination, difÞculties sometimes arise in being certain if the scleral disease with or without corneal involvement is completely under control. Anterior segment ßuorescein angiography can sometimes be of assistance in monitoring the effect of medical therapy [85].

If in spite of intensive medical therapy no new vessels are formed or no preexisting vessels are recanalized, progressive thinning of the sclera and/or cornea with possible eventual perforation may occur. In these cases, tectonic surgery, such as scleral, corneal, or sclerocorneal grafting, must be considered to maintain the integrity of the eye [86]. Although the site and extent of the surgical procedure can be frequently decided by clinical examination, determination of the amount of necrotic tissue to be removed and replaced surgically can sometimes be difÞcult. Anterior seg-

ment ßuorescein angiography can sometimes be useful in deciding the extent of appropriate surgical intervention [85].

3.2.6.1 Anterior Segment Fluorescein Angiography Techniques

Conventional photographic ßuorescein angiography [81, 87Ð93], low-dose photographic ßuorescein angiography [94], low-dose ßuorescein videoangiography [95], and scanning angiographic microscopic ßuorescein videoangiography [96] are different techniques used to describe the circulatory dynamics of the anterior segment of the eye, such as the direction of ßow, distinction between arteries and veins, and integrity of the circulation.

Fluorescein angiography has been available for examining the retinal microcirculation since 1961 [97], when venous injection of low-molec- ular-weight sodium ßuorescein was used to demonstrate abnormalities in the retinal capillaries, retinal pigment epithelial cells, and BruchÕs membrane. The tight apposition of contiguous retinal capillary endothelial cells may explain, at least in part, why normal retinal vessels do not leak ßuorescein [88, 98]. Iris capillary endothelial cells also are joined by tight junctions, and anterior segment ßuorescein angiography was introduced in 1968 [99, 100] with the primary purpose of diagnosing iris lesions. Conventional anterior segment ßuorescein angiography has not, however, been widely used for conjunctival and scleral abnormalities because normal conjunctival and episcleral vessels leak molecules smaller than serum albumin, such as ßuorescein. Low-molecular-weight molecules may escape from the conjunctival and episcleral vessel lumens by crossing the interendothelial clefts, endothelial cells through pinocytotic vesicles, or both [101]. Interestingly, the limbal vessels never leak ßuorescein, suggesting that their endothelial cells are united by tight junctions [102]. Five milliliters of 10% sodium ßuorescein via antecubital vein injection rapidly extravasates from conjunctival and episcleral vessels, restricting the diagnostic value of this technique to the demonstration of either early leakage or gross hypoperfusion [81]. If leakage is to be avoided, the ßuorescein