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PubMed2001

Drug delivery in poly(lactide-co-glycolide) nanoparticles surface modified with poloxamer 407 and poloxamine 908: in vitro characterisation and in vivo evaluation.

Poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles of 150-nm mean size were produced by an interfacial deposition method. The polar model drug Rose Bengal was successfully loaded into the nanoparticles during production and the surface of these particles was subsequently modified with poloxamer 407 and poloxamine 908 in order to create a steric stabilising layer of PEG on the surface. Drug loading was low (<1%) which can be attributed to the polar nature of the drug and the small size of the nanoparticles. Drug release was biphasic with 50% release measured within 30 min in serum. After intravenous injection in rats, the drug loaded nanoparticles substantially avoided capture by the Kupffer cells of the liver as compared to free drug. The half-life of Rose Bengal in the blood stream when administered in the nanoparticles was greatly extended with approximately 30% remaining after 1 h as compared to only 8% of Rose Bengal left 5 min after administration in solution. These surface modified nanoparticles would have potential as carriers for drugs to specific sites within the body or for slow release of drug within the circulation.

Animals↗
PubMed1981

Glutathione S-transferases in elasmobranch liver. Molecular heterogeneity, catalytic and binding properties, and purification.

In order to gain insight into the phylogeny and physiological significance of organic-anion-binding proteins in the liver, the hepatic glutathione S-transferases of rat and a typical elasmobranch, the thorny-back shark (Platyrhinoides triseriata), were compared with respect to both glutathione S-transferase activites and organic-anion-binding properties. On gel filtration (Sephadex G-75, Superfine grade) of rat cytosol, the elution volumes of enzyme activities with 1-chloro-2,4-dinitrobenzene and p-nitrobenzyl chloride as substrates were identical (rat Y-fractions; M(r) 45000). In contrast, two peaks of enzyme activity for 1-chloro-2,4-dinitrobenzene with elution volumes corresponding to M(r) 52000 (PLAT Y(1)) and M(r) 45000 (PLAT Y(2)) were detected on gel filtration of P. triseriata cytosol. Only fraction PLAT Y(2) had enzyme activity with p-nitrobenzyl chloride. Enzyme kinetic studies showed that rat Y-fraction had higher affinities for both 1-chloro-2,4-dinitrobenzene and glutathione than PLAT Y(1)- and PLAT Y(2)-fractions. The two forms of P. triseriata glutathione S-transferases differed greatly in affinity for glutathione. At a glutathione concentration that we found to be physiological in P. triseriata, PLAT Y(2) accounted for approx. 70% of the total glutathione S-transferase activity with 1-chloro-2,4-dinitrobenzene. Binding studies revealed that PLAT Y(1) and PLAT Y(2) fractions had much lower affinities for sulphobromophthalein and bilirubin than rat Y-fraction. In contrast, binding affinities of PLAT Y(1) and PLAT Y(2) for Rose Bengal and 1-anilino-8-naphthalenesulphonate were comparable with that of rat Y-fraction. Inhibitory kinetics suggested that sulphobromophthalein and Rose Bengal were non-competitive inhibitors of glutathione S-transferase activities when 1-chloro-2,4-dinitrobenzene was used as substrate for both PLAT Y(1) and PLAT Y(2). The major glutathione S-transferase from the PLAT Y(2) fraction was purified 81-fold by sequential chromatography on Sephadex G-75, DEAE-Sephadex and hydroxyapatite, and consisted of two identical subunits with pI7.7. The highly enriched Y(2)-fraction retained high affinity binding of Rose Bengal and 1-anilino-8-naphthalenesulphonate.

Amino Acids↗
PubMed1975

Some serological reactions to "brucella" antigen in the horse.

Seventy-three samples of serum, from 69 horses and one zebra, were subjected to the Rose Bengal Plate, serum tube agglutination, complement fixation, and anti-equine globulin (Coombs') tests for brucellosis. Fifty-one of the samples, from 48 horses, were submitted by practising veterinary surgeons; of these, 22 samples were associated with clinical conditions which might have been due to brucellosis. Fourteen samples were from healthy horses known to have been in contact with infected cattle, and six were from horses which were known not to have been exposed to brucellosis. More reactions at accepted diagnostic levels were obtained to the anti-globulin and complement fixation tests than to the agglutination test. Two horses showed no reactions, other than inconclusive titres to the antiglobulin test, and these titres could have resulted from exposure to Brucella. Eight of nine positive Rose Bengal tests were confirmed by a reaction at diagnostic level in at least one of the other tests, but two sera showing a reaction at diagnostic level to the other tests gave a negative Rose Bengal result.

Agglutination Tests↗
PubMed1988

[Observation of the acrosome reaction of human spermatozoa using the combined preparation with triple stain technique and scanning electron microscopy].

The combination of triple stain technique (TST) and scanning electron microscopy (SEM) was used to investigate the reliability of TST in observing the acrosome reaction. The TST preparation was followed by SEM to observe the same spermatozoa by both methods. We compared the results obtained with TST at various pH of rose Bengal solution to those with SEM. 1. In our method, the acrosome reaction on the same spermatozoa was easily examined by both TST and SEM. 2. The fine surface structure of the sperm head was not affected by the TST procedure, because the percentages of acrosome non-reacted (ANR) and acrosome reacted (AR) sperm were similar when spermatozoa from the same semen were examined by SEM or SEM after TST (23.0%, 77.0% VS. 24.5%, 75.5%, respectively). 3. When results obtained with TST at various pH of rose Bengal solution were compared to those of SEM after TST, the accuracy was 70.5% at pH 4.3, 95.9% at pH 5.3 and 61.8% at pH 6.3. Sensitivity and specificity were 100%, 21.4% at pH 4.3, 94.5%, 97.6% at pH 5.3 and 38.7%, 97.5% at pH 6.3 respectively. From these results, it can be concluded that TST with pH 5.3 rose Bengal solution is a reliable and practical method for detecting the acrosome reaction of human spermatozoa.

Acrosome↗
PubMed1995

Oxidative reactions in the tear fluid of patients suffering from dry eyes.

PURPOSE: To evaluate whether products of oxidative and inflammatory reactions are detectable in the tear fluid of patients suffering from dry eyes. METHODS: The tear fluid of 217 patients (397 eyes) was sampled. Criteria for grouping of the patients were (1) basic secretion test ("sicca 1": BST = 0-5 mm, n = 78 eyes; "sicca 2": BST = 6-10 mm, n = 109 eyes) and (2) subjective symptoms (normal BST, burning, foreign body sensations, tearing, dryness of the eyes: n = 78 eyes). One group of healthy patients (normal BST, n = 132 eyes) served as controls. Lipid peroxide levels and myeloperoxidase activity, as parameters for oxidative tissue damage and inflammatory activity, were determined in the tear fluid. Those patients whose consent could be obtained were subjected to the rose bengal test (sicca 1: 56 eyes; sicca 2: 97 eyes; subjective symptoms: 44 eyes; controls: 49 eyes). The correlation between BST and rose bengal test results was calculated. RESULTS: Lipid peroxides were significantly (P < 0.05) higher in the groups sicca 1 and subjective symptoms than in healthy controls, as was the inflammatory activity in groups sicca 1, sicca 2 and subjective symptoms. Additionally, the inflammatory activity in the group sicca 1 was significantly (P < 0.05) higher than in the groups sicca 2 and subjective symptoms. No evidence of a significant correlation between BST and rose bengal test results was observed. CONCLUSIONS: Both oxidative tissue damage and polymorphonuclear leukocytes indicating an oxidative potential occur in the tear film of patients suffering from dry eyes. These reactions lead to severe damage of the involved tissue. Free radicals and inflammation may be involved in the pathogenesis or in the self-propagation of the disease.

Dry Eye Syndromes↗
PubMed1986

Rh antigen immunoreactivity after histidine modification.

125I-anti-D IgG and unlabeled blood group allo-antisera in combination with 125I-protein A were employed in assessing antibody binding to red cells (RBC) treated with histidine reagents. The acylating reagent diethylpyrocarbonate (DEP), the alkylating reagent p-bromophenacyl bromide (pBPB) and the photosensitizer dye Rose Bengal (RB) were used under conditions that usually result in the selective modification of histidine in isolated proteins. Progressive apparent inactivation of the D antigen in ghost membranes occurred with increasing DEP concns, which was not demonstrably reversible by hydroxylamine since this reagent itself inactivated the D antigen. Exposure of red cells to 5 mM p BPB resulted in a 50% decrease in binding of 125I-anti-D IgG. Photo-oxidation of RBC in the presence of Rose Bengal apparently inactivated all the major Rh antigens as detected either by labeled anti-D IgG binding, IgG agglutinating serological reagents, or the binding of 125I-labeled protein A following the sensitization of cells with unlabeled antisera. Under conditions of RB treatment, where hemolysis was absent or minimal, 125I anti-D IgG binding decreased to 38-49% of the level seen in controls. Rose Bengal treatment of R1r RBC revealed varying inactivation of all the Rh antigens, i.e. D 15%, C 89%, c 73%, e 54% inactivated, whereas antibody binding activity of the Fya and Fyb antigens present in the same cell was unaffected. Previous reports as well as the pH profile of anti-D binding have implicated the participation of histidine in Rh antigen expression. Our results are consistent with histidine involvement in Rh activity. Whether Rh antigens have essential histidine(s) involved directly in epitope structure, or instead depend on a critical histidine(s) at the lipid-protein interface that modulates antigen expression remains to be determined.

Diethyl Pyrocarbonate↗
PubMed2005

[Comparison of the serological tests used for the laboratory diagnosis of brucellosis].

Brucellosis which is an important public health problem is a zoonotic disease that causes economic loss, and seen all over the world as well as in our country. The aim of this study was to compare the results of Rose Bengal test, standard tube agglutination test (STA), Coombs tube agglutination test, Rivanol tube agglutination test and ELISA (IgA, IgG and IgM) method, in patients who were suspected to have brucellosis. Blood and serum samples collected from 77 patients were included to the study. Blood cultures were performed from all of the patients and Brucella spp were isolated in 35 (45.4%) of them. At least one of the serological tests for Brucella was found positive in 62 samples (80.5%). When culture positivity accepted as the gold standard, the sensitivities of the tests were calculated as follows; 100% for Rose Bengal, 94.3% for STA, 97.1% for ELISA IgG, 94.3 for ELISA IgA and 71.4 for ELISA IgM. No prezone phenomenon was observed in the samples. Brucella Coombs tube agglutination test did not increase the chance of diagnosis of brucellosis, unlike the results of previous studies. We found that Rose Bengal and STA tests were still efficient methods for brucellosis serodiagnosis. The results of rivanol tube agglutination test were similar to STA test, and this test can be used especially in patients with brucellosis relapse. ELISA test detecting IgA and IgG antibody titers together found to be effective methods increasing the chance of brucellosis diagnosis, and could be helpful for follow up of brucellosis.

Agglutination Tests↗
PubMed2003

Corneal surface changes in keratoconjunctivitis sicca. Part I: The surface proper. A non-contact photomicrographic in vivo study in the human cornea.

PURPOSE: To describe corneal surface changes in keratoconjunctivitis sicca (KCS). METHODS: In all 23 patients with KCS were examined with the slit lamp and photographed by non-contact photomicrography. RESULTS: Without staining, grey surface cells and small epithelial cysts were visible. Application of fluorescein sodium resulted in two types of micropunctuate (15-30 microm) staining, a nonfluorescent (yellow/brown) and a fluorescent (green) one, in brilliantly green staining of cysts, and in enlarging fluorescent (green) flecks (70-1600 microm) with propensity to confluence. The flecks often developed in inconspicuous areas and showed abnormal subsurface cells. Application of rose bengal resulted in more or less intensively red-stained cells measuring about 30 mm in diameter, often less (15-20 microm), with or without a centrally located nucleus, individual or gathered in small groups or larger patches, and in confluent staining. Smaller and larger cells, with or without nuclei, weakly or strongly stained, were often located close to each other. The smaller green flecks showed a few centrally located red-stained cells, and the larger ones showed many almost uniform small red-stained nucleated ones. CONCLUSIONS: Corneal epithelial surface disease in the present KCS patients consisted of unspecific features-such as increased light-reflecting property (grey cells), disruptions of intercellular junctions (micropunctuate green fluorescein staining), pathological dye uptake (micropunctuate yellow/brown fluorescein and red rose bengal stainings), and epithelial oedema (microcysts)--and also of features that seemed to be disease specific. Such features were, with fluorescein sodium, focal disruptions of the epithelial barrier function manifesting in enlarging green flecks, and, with rose bengal, abundant nucleated cells, particularly small ones organised in foci. The nature of the abnormal subsurface cells present in the areas of the green flecks, and the mechanisms behind the focal involvement are unclear.

Adult↗
PubMed2006

Clinical evaluation of the Smart Plug in the treatment of dry eyes.

PURPOSE: To evaluate the efficacy of a thermo sensitive punctum plug, the Smart Plug, in the treatment of dry eyes. DESIGN: Observational case-series study. METHODS: Eighteen eyes of 10 dry eye patients who previously failed conventional plug insertion were enrolled. Schirmer 1 test, vital staining scores, and tear clearance tests were performed before plug insertion, at 2 weeks, 1 month, and 3 months after insertion. RESULTS: Although Schirmer test values were not significantly different before and after plug insertion, the tear clearance rate considerably decreased (Before insertion: 14.8+/-11.4; After insertion: 8.2+/-6.6) (P<.05). Rose-Bengal and fluorescein scores improved significantly after plug insertion (Rose-Bengal: Before insertion: 6.4+/-2.0; After insertion: 3.3+/-1.6 points, Fluorescein: Before insertion: 4.8+/-2.3; After insertion: 2.1+/-1.3) (P<.05). No complications were observed. CONCLUSIONS: Smart Plug proved to be a safe and efficient option in the treatment of dry eye.

Dry Eye Syndromes↗
PubMed2005

Staining properties of deepithelialized human amniotic membrane.

PURPOSE: The surgical use of human amniotic membrane (HAM) can be challenging because of its transparency, thin structure, and adhesive quality. Staining of HAM has been proposed to facilitate intraoperative visualization. We evaluated the in vitro staining properties of deepithelialized HAM using 5 vital dyes. METHODS: Deepithelialized HAM was stained with indocyanine green (1.0%, 0.5%, 0.1%), fluorescein sodium (1.0%, 0.25%, 0.1%), lissamine green (0.5%, 0.1%), rose bengal (1.0%, 0.1%), and trypan blue (0.5%, 0.1%). The staining of each dyed sample was evaluated using a x10 operating microscope by a single observer. Each membrane was then rinsed, resuspended (3 mL of balanced salt solution), and then reevaluated for staining at 30-minute intervals for a total of 4 hours. All remaining stained samples were then placed in 5 mL balanced salt solution and then reevaluated for staining at 24 hours. RESULTS: All concentrations of the 5 dyes stained the membrane initially. After 120 minutes, fluorescein sodium 0.1% and lissamine green 0.5% and 0.1% no longer stained the membrane. Fluorescein sodium 0.5% no longer stained at 210 minutes, and fluorescein sodium 1.0% no longer stained at 24 hours. All concentrations of indocyanine green, rose bengal, and trypan blue stained positively at 24 hours. CONCLUSIONS: All 5 dyes stain deepithelialized HAM initially. Tested concentrations of fluorescein sodium and lissamine green may be superior to other tested dyes for intraoperative use because these dyes stain the membrane and then fade. Tested concentrations of rose bengal, trypan blue, and ICG may not be ideal for clinical use because they demonstrate persistent staining at 24 hours.

Amnion↗
PubMed1992

Adsorption of fluorescein dyes on albumin microspheres.

The surface characteristics of bovine and egg albumin microspheres were examined using four anionic dyes; sodium fluorescein, eosin, erythrosin, rose bengal, and the cationic dye rhodamine B. The adsorption isotherms of the dyes on unloaded albumin microspheres exhibited Langmuir behavior for dilute solutions of rose bengal, erythrosin, and eosin, suggesting monolayer formation in the initial stages of the sorption process. The adsorption capacity of the microspheres for the dyes (k2) and the affinity constants of the dyes for the microspheres (k1) were found to depend on both the polarizability and the hydrophobic properties of the dye, presumably reflecting the heterogeneous character of the microsphere surface. Further, the extent of sorption at higher dye concentrations was found to depend on the ability of the dye to form stable aggregates inside the microspheres and on environmental long-range forces acting at these sites. At both low and high dye concentrations, the amount adsorbed to the microsphere surface increased with increasing hydrophobicity of the dyes. The lowest adsorption was observed for the nonsubstituted dye fluorescein, whereas the most hydrophobic dye used, rose bengal, was completely adsorbed onto the microsphere surface. The data suggest that the bovine albumin microsphere surfaces are highly hydrophobic and less porous than egg albumin microsphere surfaces.

Adsorption↗
PubMed1998

The epidemiology of dry eye in Melbourne, Australia.

OBJECTIVE: To describe the epidemiology of dry eye in the adult population of Melbourne, Australia. DESIGN: A cross-sectional prevalence study. PARTICIPANTS: Participants were recruited by a household census from two of nine clusters of the Melbourne Visual Impairment Project, a population-based study of age-related eye disease in the 40 and older age group of Melbourne, Australia. Nine hundred and twenty-six (82.3% of eligible) people participated; 433 (46.8%) were male. They ranged in age from 40 to 97 years, with a mean of 59.2 years. MAIN OUTCOME MEASURES: Self-reported symptoms of dry eye were elicited by an interviewer-administered questionnaire. Four objective assessments of dry eye were made: Schirmer's test, tear film breakup time, rose bengal staining, and fluorescein corneal staining. A standardized clinical slit-lamp examination was performed on all participants. Dry eye for the individual signs or symptoms was defined as: rose bengal > 3, Schirmers < 8, tear film breakup time < 8, > 1/3 fluorescein staining, and severe symptoms (3 on a scale of 0 to 3). RESULTS: Dry eye was diagnosed as follows: 10.8% by rose bengal, 16.3% by Schirmer's test, 8.6% by tear film breakup time, 1.5% by fluorescein staining, 7.4% with two or more signs, and 5.5% with any severe symptom not attributed to hay fever. Women were more likely to report severe symptoms of dry eye (odds ratio [OR] = 1.85; 95% confidence limits [CL] = 1.01, 3.41). Risk factors for two or more signs of dry eye include age (OR = 1.04; 95% CL = 1.01, 1.06), and self-report of arthritis (OR = 3.27; 95% CL = 1.74, 6.17). These results were not changed after excluding the 21 people (2.27%) who wore contact lenses. CONCLUSIONS: These are the first reported population-based data of dry eye in Australia. The prevalence of dry eye varies by sign and symptom.

Adult↗
PubMed1999

Diseases associated with ocular surface abnormalities: the importance of reflex tearing.

AIM: To investigate the correlation between tear function tests and ocular surface integrity in patients with dry eye. METHODS: 297 dry eye patients (55 Sjögren's syndrome, two male and 53 female, average age 52.4 (SD 15.0) years, and 242 non-Sjögren's syndrome, 41 male and 201 female, average age 53.5 (14.1) years) were examined. The following tear function tests were performed: (1) cotton thread test, (2) Schirmer test with topical anaesthesia, (3) Schirmer test without anaesthesia, (4) Schirmer test with nasal stimulation, (5) tear clearance test, and (6) tear break up time (BUT). The ocular surface was evaluated by rose bengal and fluorescein staining. Correlation analysis was performed between each tear function index and vital staining scores. RESULTS: Among the six tear function tests, the Schirmer test with nasal stimulation correlated most with both of the vital stains (rho = 0.530 for rose bengal and 0.393 for fluorescein). The Schirmer test with or without anaesthesia correlated slightly with rose bengal staining, whereas tear clearance test and tear break up time slightly correlated with fluorescein staining. CONCLUSION: Vital staining of the ocular surface correlates most with reflex tearing measured by the Schirmer test with nasal stimulation.

Dry Eye Syndromes↗