Zemira Samra

Proteins in tears from healthy and diseased eyes

The levels of total protein in tears from healthy donors, conjunctivitis vernalis patients, and conjunctivitis follicularis patients, were 625, 1370 and 1160 mg% respectively. Serum albumin accounted for 3.3%, 43% and 67% of the total protein of tears from these groups, and the level of proteins probably synthesized by the lacrimal gland, was in tears from conjunctivitis follicularis patients only half the level in normal tears. By crossed immunoelectrophoresis with intermediate gel, 10 antigenic species could be recognized in normal tears, and of these the following were identified: Lysozyme, IgA, lactoferrin and serum abumin. In tears from patients with conjuntivitis vernalis three more immunoprecipitates were observed, of which one was due to IgG. No lysozyme could be demonstrated in tears from a case of conjunctivitis sicca by immunoelectrophoresis.In tears from healthy donors the mean level of IgA was 20 mg%, of IgG 3 mg%, and IgM could not be demonstrated. Rabbit anti-tear immunoglobulin did not precipitate a standard of human IgM in double immunodiffusion. In cases of conjunctivitis vernalis and follicularis the mean levels were increased to 80 and 114 mg% IgG, and 11 and 14 mg% IgM, but IgA was increased only to 32 and 41 mg%. It is assumed that the level of IgA in normal tears is almost entirely due to local synthesis, while serum albumin and other immunoglobulins may have escaped from the circulation by molecular sieving. The increased levels of immunoglobulins in inflammatory diseases is probably due to transudation. However, in blepharoconjunctivitis patients several tear samples with a high IgM and a low or zero level of IgG could be demonstrated. Possible explanations for this phenomenon are discussed.

An antigen common to a wide range of bacteria. 2. A biochemical study of a "common antigen" from Pseudomonas aeruginosa.

Common Antigen (CA) of Pseudomonas aeruginosa has been shown to be a protein composed of polypeptide subunits of a molecular weight (MW) of about 62 000. The MW of this protein was estimated to 665 000 by gel filtration on sepharose CL-6B, to 800 000 by electrophoresis on polyacrylamide gradient gels and to about 900 000 by ultracentrifugation, on a sucrose gradient. By analytical ultracentrifugation with Schlieren optics a sedimentation coefficient (S20 degrees, W) of 22.65 was calculated. The isoelectrical point was determined to pH 4.4. The antigen was decomposed on exposure to proteolytic enzymes. Polysaccharide, lipid, deoxyribonucleic acid or ribonucleic acid were not demonstrated in CA. The amino acid content of CA was determined, and no hexosamine or abnormal residues were observed. The amino acid content of CA was determined, and no hexosamine or abnormal residues were observed. The antigen was degraded when heated to 100 degrees C for 4 min or when exposed to pH below 4 or above 11 at 4 degree C. CA has been isolated from the cytoplasmic water-soluble fraction of disintegrated bacteria and only trace-amounts could be obtained from envelope fractions after solubilization with Triton X-100.

Allergen-Specific Immunoglobulin E Antibodies in Tears and Serum of Vernal Conjunctivitis Patients

Tears and sera of 53 patients of vernal conjunctivitis were examined for antibodies of the IgE type to a panel of 18 allergens. In 18 of the patients (34.0%) allergen-specific IgE was demonstrated in both tears and serum, in 3 (5.7%) in tears only, and in 4 (7.5%) in serum only. The antigen-specific reactions with tear fluid were found in patients with the highest total IgE levels, not only in tears but also in serum. This is evidence for a specific, local allergic reaction in these patients. Most positive reactions were to perennial allergens, particularly house dust mites, cat epithelium and Bermuda grass. This fact is in harmony with the lack of season-linked symptoms in most patients in this geographical area.

An immunological study of papillary conjunctivitis due to contact lenses

Serum and tear levels of IgE were compared in patients with contact lens papillary conjunctivitis (CLPC), vernal keratoconjunctivitis (VC), healthy contact lens wearers (CLW), and healthy controls without lenses. The mean of serum IgE was elevated only in the VC group. Tear IgE levels were significantly higher for CLPC and VC than for CLW and healthy controls. Tear IgG levels were also increased in CLPC patients. With serum albumin (HSA) as a marker for leakage of proteins from the circulation to the tear fluid, the data indicated, that tear IgG was blood-borne whereas tear IgE was essentially a product of local synthesis. In one of the 10 CLPC patients, high titers of IgE type antibodies to housedust mites and cat epithelium were demonstrated in both serum and tears. We conclude that CLPC is usually an IgE mediated reaction to the lens material or to contaminations of allergenic material that sticks to the lenses.

Vernal Keratoconjunctivitis: The Significance of Immunoglobulin E Levels in Tears and Serum

Vernal keratoconjunctivitis (VC) is an inflammatory disorder of the outer eye, probably of allergic etiology. The present study was performed on patients with a typical clinical appearance of VC and examined periodically for serum and tear IgE and albumin (HSA). The geometric mean of serum and tear IgE was 219 and 18 IU/ml, respectively, for the patients, but about 100 and less than 1 IU/ml for serum and tears of healthy controls and of parents of VC patients. A positive correlation between variations in tear IgE and tear HSA was observed, indicating that tear IgE levels have a significant influence on the intensity of the inflammatory process.

A contribution to the immunopathology of vernal keratoconjunctivitis

The tear fluid contains proteins which are synthesized locally in the conjunctiva and lacrimal glands, and others, which reach the tear fluid from the blood circulation. It is generally accepted that serum albumin (HSA) belongs to the latter group of tear proteins. Consequently, the ratio between the levels of HSA in the tear fluid and in the blood serum can be used as a parameter for the degree of vascular leakage. All blood-borne tear proteins will show concentration ratios in tears and blood serum similar to HSA, provided that they pass through the blood-tear barrier at the same flow rate as HSA. This principle may be used to determine whether a certain tear protein is blood-borne or locally synthesized. An additional method to determine the source of tear proteins is supposed, and is based on the assumption that blood-borne proteins will occur in tears from both eyes at a concentration ratio similar to that of HSA, whereas the concentration ratios of locally synthesized proteins will not be related to the HSA-levels in the tears of the two eyes. This method is independent of the rate of diffusion through tissue barriers.In the present study these methods have given coordinating and supplementary information. The IgG and IgM contents of tears from healthy eyes and from patients with vernal keratoconjunctivitis (VC) were shown to be essentially blood-borne. In tears from VC-patients as well as from healthy controls, IgE was usually found to be a product of local synthesis. In tears from healthy eyes, IgA was almost entirely synthesized locally, but a significant part of the IgA in tears from inflamed eyes was blood-borne.The geometrical mean (GM) of blood levels of IgE for VC-patients was not increased in relation to the GM for healthy controls of the same age groups. In contrast, the GM for IgE in tears from VC-patients (27.5 I.U./ml) was higher than the GM + 2 SD for the healthy controls (4.1 I.U./ml) and also than the highest level of IgE observed in tears from healthy eyes (6.6 I.U./ml). The tear-levels of IgE for VC-patients were strikingly dynamic and in some patients a hundred-fold increase or decrease was observed in the course of a relatively short time period. In contrast, alterations of blood-levels of IgE and of tear levels of IgA were relatively moderate. These results indicate a marked association between VC and an increased IgE-content of tears derived from local immunocytes. However, in some patients apparently suffering from typical VC, normal IgE-levels were consistently found in tears from both eyes during a number of relapses or aggravations of the disease. These results may indicate that more than one etiopathogenesis may lead to the clinical condition known as VC. A hitherto unexplained feature of VC is the pronounced lacrimation. We have shown that the concentration of the essential tear-proteins is not decreased in VC. Lacrimation is therefore associated with accelerated protein synthesis of the secretory system.

A Sensitive and Wide-Scale Enzyme-Immunoassay for Human Transferrin

We describe an ELISA technique for human transferrin. The test has a lowest detection limit of 2 micrograms/l and a wide range. It is specific for human transferrin and its reproducibility proved acceptable in both intra-run and between-day examinations. Results obtained by radial immunodiffusion and by ELISA were strongly correlated (n = 51; y = 0.92x + 16.1; r = 0.90). The cost in Israel for disposables and chemicals for triplicate examination is

Immunoglobulins of the Subretinal Fluid

0.55 per sample. This is 7-8 times less than the cost of a determination, using commercial immunodiffusion plates. This assay is of particular value where the sample size is limited, e.g. samples of 1-4 microliters are sufficient for the determination of transferrin in tears. The transferrin concentrations in arterial and venous cord blood were shown to be essentially identical and similar to that in adult blood.

An antigen common to a wide range of bacteria.I. The isolation of a "common antigen" from Pseudomonas aeruginosa.

The formation of subretinal fluid (SRF) in rhegmatogenous retinal detachment is a complex process involving the vitreous (Cooper, Halbert & Manski 1963), the sensory retina and retinal pigment epithelium (Leopold & Furman 1971; Feeney, Burns & Nixon 1975) and the choriocapillaris (Kaufman & Podos 1973). It is generally agreed that the greatest part of the protein content of the SRF derives from the choroid (Liotet & Rouchy 1972; Chignell, Carrhuters & Rahi 1971) and that the choriocapillaris-retinal pigment epithelium barrier is an effective barrier, the permeability of which increases with the duration of the retinal detachment (Chignell, Carrhuters & Rahi 1971). It has been stated that, not the total protein content of the SRF but rather the differential protein study is a good indicator of the damage of this barrier (Chignell, Carrhuters & Rahi 1971; Rahi & Chignell 1975).