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PubMed1981

Preparation, specific binding, and biological activity of a monosubstituted fluorescein derivative of prolactin.

A monosubstituted fluorescein derivative of ovine prolactin has been prepared by the reaction of fluorescein isothiocyanate with the hormone at pH 8.0. Reaction under the specified conditions results in a derivative incorporating one fluorescein moiety per molecule of prolactin as judged by coordinated UV spectroscopy and amino acid analysis. The absence of NH2-terminal threonine in the derivative indicates that the point of fluorescein attachment is the NH2-terminal amino group, whose substitution is expected to be favored at pH 8.0. The derivative was found to retain significant specific binding on both rabbit mammary and rat ventral prostate membrane preparations in competition with 125I-labeled prolactin, and to show saturable stimulation of progesterone production in primary cultures of rat granulosa cells. It is anticipated that this derivative will be useful in studies of prolactin-target cell interactions. This study shows that NH2-terminal monosubstituted derivatives of ovine prolactin are readily formed under specified conditions at pH 8.0; these conditions may be useful in the preparation of other derivatives of this type.

Animals↗
PubMedDate not supplied

Photoelectron microscopy and photoelectron quantum yields of the fluorescent dyes fluorescein and rhodamine.

Photoelectric properties of the dyes fluorescein and rhodamine were determined with the aim of assessing the usefulness of these compounds as labels in photoelectron microscopy. The photoelectron quantum yields were measured over the wavelength range 180-230 nm. At 230 nm the quantum yields for fluorescein disodium salt, rhodamine B free base and rhodamine B HCl salt are approximately 10(-5) electrons per incident photon. At 180 nm these values rise to approximately 10(-3) electrons per incident photon. All forms of fluorescein do not have the same quantum yield. The neutral form of fluorescein has a quantum yield an order of magnitude lower than the disodium salt. Beam current measurements were performed on labeled and unlabeled proteins to determine the effect of the high light intensity employed in the photoelectron microscope. The initial beam current measurements and the quantum yield curves are consistent and demonstrate that there is significant contrast between labeled and unlabeled proteins. However, after several minutes in the photoelectron microscope, the proteins become more photoemissive and the contrast diminishes. This change in contrast explains several puzzling observations in the literature.

Electrons↗
PubMed1986

Aqualuminescence of alkaline luminol in the presence of fluorescein.

The light yield from both luminol and fluorescein is studied at a fixed concentration of luminol and by varying the concentration of fluorescein in 3 X 10(-2) M basic solution by direct chemical reaction as well as by aqualuminescence technique. The emission spectra of chemiluminescence and aqualuminescence of the alkaline luminol-fluorescein mixture are recorded on a Fuoss spectrograph. A 4-fold increase in the aqualuminescence intensity of luminol has been observed in the presence of fluorescein as compared to that of pure luminol. The results are explained on the basis of reactions of colour centres with the activators.

Fluorescein↗
PubMed2005

Adenosine agonist regulation of outward active transport of fluorescein across retinal pigment epithelium in rabbits.

To investigate the effect of an adenosine agonist, 2-5'-N-ethylcarboxamidoadenosine (NECA), on the outward active transport of fluorescein across the retinal pigment epithelium (RPE) in rabbits. High (5x10(-4)-2x10(-3) M) and low (1x10(-5)-1x10(-4) M) concentrations of NECA or phosphate buffered saline (PBS) were intravitreously injected into Dutch-belted rabbits. Sodium fluorescein was injected intravenously 180 min after NECA. Differential vitreous fluorophotometry was performed 3 hr after the sodium fluorescein injection and the vitreal fluorescein/fluorescein monoglucuronide (F/FG) ratio then was calculated. The F/FG ratios are inversely proportional to the outward active transport of fluorescein across the RPE. Retinal detachments were induced by injection of PBS into the subretinal space after the intravitreous injection of low- or high-dose NECA or PBS, and the size of the blebs was monitored. In eyes that received a low-dose injection of NECA, the F/FG ratio was higher compared with controls (P<0.05); in eyes that received a high-dose intravitreal injection, the F/FG ratio was significantly lower compared with controls (P<0.05). The effect of low-dose NECA on the F/FG ratio was suppressed by the A2 receptor antagonist, ZM241385, and the effect of high-dose NECA was suppressed by the A1 receptor antagonist, 8-cyclopentyl-1, 3-dipropylxanthine. The A3 receptor antagonist MRS1191 did not influence the effect of low- or high-dose NECA. Intravitreal injection of high-dose NECA enhanced the reabsorption of subretinal fluid compared with PBS; however, low-dose NECA inhibited reabsorption of subretinal fluid (P<0.02 and 0.05, respectively). Intravitreous injection of high-dose NECA accelerates the active outward transport across the RPE via A1 receptors and low-dose NECA decelerates it via A2 receptors.

Adenosine↗
PubMed1990

Potential bacterial contamination in fluorescein-anesthetic solutions.

To determine the ability of fluorescein-anesthetic combination solutions and their applicators to regain sterility, we contaminated four commercially available fluorescein-anesthetic solutions and their dropper tips with inocula of either Pseudomonas species or Staphylococcus species. No organisms could be cultured from Fluress one minute after inoculation of the solution or five minutes after inoculation of the dropper tip. In contrast, organisms were cultured from the other fluorescein-anesthetic preparations for at least one hour after bacterial inoculation into the solution or onto the dropper tip. These differences in the ability of fluorescein-anesthetic solutions to regain sterility after bacterial contamination were statistically significant.

Anesthetics↗
PubMed2000

Hold up of dye in the arm during fluorescein angiography: a quantitative demonstration.

PURPOSE: To demonstrate a hold-up of part of the fluorescein bolus in the arm as a result of arm position. METHOD: Case report. We obtained a fluorescein angiogram with a calibrated confocal scanning laser ophthalmoscope in a 20-year-old healthy subject. During and after injection, the upper arm was held in approximately 60 degrees abduction, 65 degrees exorotation, and slight anteflexion. In the late venous phase, the subject moved the upper arm on the injected side to a more neutral, downward position. RESULTS: We measured a distinct rise in fluorescence level about 10 seconds after movement of the arm. The most likely explanation is relief of a partial obstruction of the venous drainage, which had been caused by the position of the upper arm described above. CONCLUSION: Impaired venous drainage of the injected arm caused by exorotation and abduction of the upper arm is a potentially common cause of delayed dye arrival or unexpectedly reduced contrast level during fluorescein angiography. Therefore, arm position needs attention in fluorescein angiography.

Adult↗
PubMed1985

Solute accessibility to N epsilon-fluorescein isothiocyanate-lysine-23 cobra alpha-toxin bound to the acetylcholine receptor. A consideration of the effect of rotational diffusion and orientation constraints on fluorescence quenching.

To obtain information on the disposition of alpha-toxin when bound to the acetylcholine receptor (AChR), we evaluated the accessibility of solutes to fluorescein isothiocyanate (FITC) conjugated to alpha-toxin (siamensis 3) at lysine 23 (FITC-toxin) by measuring the rate constants for iodide quenching of the fluorescence of fluorescein free in solution and FITC-toxin free in solution and bound to AChR. Relative to the free fluorescein, we observed a 55% reduction in the quenching rate constant for the unbound FITC-toxin and 80% reduction for the AChR-bound FITC-toxin. It is tempting to interpret a decrease in the quenching rate constant as due to an increase in the masking of the labeling fluorophore, which in our case would then be indicative of masking of fluorescein conjugated to the free toxin and masking of FITC-toxin, in the region of lysine 23, when bound to AChR. However, elementary considerations indicate that the quenching rate depends not only on geometrical masking factors but also on the translational and rotational mobilities of the labeled molecules as well as orientational constraints. To evaluate these effects we have established quantitative relations between the rate of fluorescence quenching, the degree of masking of fluorophore, translational and rotational rates, and orientational constraints of the labeled macromolecules, using recent formulations for the rate of reaction between asymmetric molecules (Shoup et al., 1981, Biophys. J., 36:619-714). These relations predict that the decrease in quenching constant observed for the labeled FITC-toxin as well as the AChR-bound FITC-toxin is largely due to differences in translational and rotational rates and orientational constraints and not to significant increases in geometrical masking. Our theoretical formulation shows that the quenching rate can be decreased by a factor of 2-5 merely by immobilizing a fluorophore on the surface of a large protein without any significant increase in geometrical masking.

Animals↗
PubMed2001

Temperature effect on nausea during fluorescein angiography.

PURPOSE: Nausea is the most common adverse effect of fluorescein angiography (FA). Warming of fluorescein dye before injection has been reported anecdotally to decrease nausea frequency. We examined the effect of warmed fluorescein dye on the frequency of adverse effects of FA. DESIGN: Prospective, randomized, controlled trial. PARTICIPANTS: Five hundred forty-seven angiograms performed on 394 patients; 273 were in the control group and 274 were in the study group. METHODS: Patients were alternately assigned to FA using dye at room temperature (control group) or dye prewarmed to body temperature (study group). All patients were masked to the dye temperature. After FA, patients were questioned regarding any adverse effects experienced. MAIN OUTCOME MEASURES: Presence or absence of reported nausea associated with FA. RESULTS: Adverse reactions were reported for 39 of the angiograms (7.1%), including 32 procedures for which nausea was reported (5.9%). No significant difference in the frequency of adverse reactions was found between the two groups (P = 0.41). No significant difference in the frequency of nausea was found between the two groups (P = 0.28). The sample sizes used allow an 80% power for detecting a clinically significant difference of 4% adverse effects in one group and 10% in the other group when testing at a significance level of 5%. CONCLUSIONS: Warming of fluorescein dye does not significantly alter the incidence of nausea reported with FA.

Female↗
PubMed1990

Delivery of fluorescein to the anterior chamber using the corneal collagen shield.

Collagen corneal shields show promise as an alternative method of drug delivery to the eye. The authors quantified collagen shield delivery of sodium fluorescein to the aqueous humor in human volunteers using fluorophotometry. Collagen shields that had been immersed in 0.01% sodium fluorescein were applied to eyes of human volunteers. The shields delivered significantly more fluorescein to the aqueous humor at 2 and 4 hours compared with drops of the same concentration instilled every 30 minutes for 4 hours (P less than 0.0001 and P = 0.0003, respectively) or daily wear soft contact lenses presoaked in 0.01% fluorescein (P less than 0.0001 and P less than 0.0025, respectively). Collagen shields did not induce damage to the corneal epithelium over a 2-hour wearing period. These results suggest that the collagen shield may be superior to topical drops in delivering water-soluble compounds to the cornea and aqueous humor.

Administration, Topical↗
PubMed1990

Urine fluorescence using a Wood's lamp to detect the antifreeze additive sodium fluorescein: a qualitative adjunctive test in suspected ethylene glycol ingestions.

Antifreeze ingestions require rapid and accurate differential diagnosis to prevent fatal outcomes. Sodium fluorescein is added to some commercial antifreeze preparations (ethylene glycol) to a final concentration of approximately 20 micrograms/mL as a colorant to aid in detection of automobile cooling-system leaks. For an adult human being, a potentially toxic volume of antifreeze is 30 mL, which contains 0.4 to 0.6 mg sodium fluorescein. Six male volunteers were given a 0.6-mg oral bolus of sodium fluorescein on an empty stomach. Urine was collected at two-hour intervals. Using a Wood's lamp, visually detectable fluorescence was seen with 100% reliability for two hours and 60% reliability for four hours. A second group of male volunteers was given the same dose of sodium fluorescein, and fluorescence was measured with a fluorometer during a six-hour period. Detectable fluorescence was present in all samples except the zero time point, including those with no fluorescence present by visual examination. We conclude that exposing urine to a Wood's lamp may be a useful adjunctive diagnostic test for early evaluation of patients with suspected antifreeze ingestion while awaiting definitive quantitative analysis of serum ethylene glycol concentration. A prospective clinical trial is needed to evaluate the frequency of false-positives and false-negatives.

Adult↗
PubMed2001

Cystoid macular edema after pediatric intraocular lens implantation: fluorescein angioscopy results and literature review.

PURPOSE: To evaluate the occurrence of cystoid macular edema (CME) after lens extraction, anterior vitrectomy, and intraocular lens implantation (IOL) in children using angioscopy after administration of oral fluorescein. SETTING: Centers in Tamil Nadu, India, and Hong Kong, China. METHODS: This study comprised 18 children (28 eyes) who had cataract extraction, posterior capsulorhexis, anterior vitrectomy, and in-the-bag IOL implantation. The presence of CME was evaluated 1 week and 4 to 6 weeks after surgery using fluorescein angioscopy. RESULTS: Anterior chamber fibrin occurred in 4 eyes (14.3%). Fluorescein angioscopy was performed 1 week after surgery in all eyes and after 1 month in 25 eyes (89.3%). No eye demonstrated the presence of CME on fluorescein angioscopy. CONCLUSIONS: Cystoid macular edema did not occur in the early period after pediatric cataract surgery using current surgical techniques. Longer follow-up is required to ascertain the incidence of CME in the late postoperative period.

Capsulorhexis↗
PubMed2000

Clinical evaluation of 'local contrast enhancement' for oral fluorescein angiograms.

PURPOSE: To describe and assess the clinical impact of 'local contrast enhancement' on oral fluorescein angiograms. METHODS: Oral fluorescein angiograms (OFA) were performed on 12 patients with a range of diabetic retinopathy. The digital images were processed using two enhancement techniques: histogram equalization, a widely available method; and 'local contrast enhancement'. Twelve control images and 24 enhanced images were randomised and subjectively graded for clarity between 1 and 100 on a visual analogue scale by two masked observers. A score of 50% was chosen as the cut-off for clinically useful images. RESULTS: The mean score for the unenhanced images was 38.8% (SD 19.4); the histogram equalization performed better at 54.7% (SD 10.0) (p = 0.016) and 'local contrast enhancement' at 69.4% (SD 13.6) (p < 0.001). These results equate to a 14.7% improvement in comparison with histogram equalization and a 30.3% improvement with the 'local contrast enhancement' technique. All the 'local contrast enhancement' images were clinically useful, compared with 58.3% of the histogram equalization images (chi 2 2.08, p = 0.2), and 33.3% of the control images (chi 2 = 6.75, p = 0.01). CONCLUSION: This pilot study has demonstrated the power of 'local contrast enhancement' techniques for low-contrast fluorescein angiograms. Using 'local contrast enhancement' oral fluorescein angiograms can give high-quality images and may allow safer patient investigation.

Administration, Oral↗
PubMed2001

Use of oral fluorescein angiography in the diagnosis of macular oedema within a diabetic retinopathy screening programme.

AIMS: To assess if oral fluorescein angiography (OFA) is a suitable screening method to detect macular oedema in diabetic retinopathy. METHODS: Eighty-four diabetic patients were included in the study. They were from a consecutive series of patients attending the diabetic eye-screening clinic, with retinopathy at the macula requiring ophthalmology assessment. All patients were subsequently examined in the eye hospital, by ophthalmologist slit lamp biomicroscopy assessment as the gold standard, followed by oral fluorescein angiography. RESULTS: This study indicates a sensitivity of 92% and specificity of 81%. Only 4.8% of patients developed a minor reaction to oral fluorescein; 84.5% of images were of good quality. CONCLUSIONS: Oral fluorescein angiography is an efficient and highly sensitive tool for the detection of macular oedema. It can be used as an adjunct in the diabetic screening service to identify patients with oedema within a disc diameter of the macula. Ultimately it will ensure that only necessary and smaller numbers of patients are referred to ophthalmologists.

Administration, Oral↗
PubMed2000

[Aqueous outflow pathway imaging in the rabbit with fluorescein and indocyanine green].

PURPOSE: The purpose of this study was to visualise the rabbit aqueous outflow pathway using a numeric imaging system with ICG and fluorescein injection in the anterior chamber. MATERIAL AND METHODS: We performed a simultaneous injection of Indocyanin Green (ICG) and Fluorescein into the anterior chamber of rabbit eyes. Using a digital camera, we took several sequenced pictures to visualize the distribution of the dyes within the outflow pathway. We observed the dynamics of the outflow over time. RESULTS: In the early phases, the shape of the outflow canal around the limbus was clearly seen. Several collecting veins close to the recti muscles were also identified. There was only slight fluorescein leakage during the early phases, allowing adequate visualization of the morphology of the outflow system. In the late phases, the sclera was stained with the fluorescein, and no details were thus visible. The ICG dye allowed better recognition of the fine details of the outflow structure. CONCLUSION: This method was relatively simple, safe and precise, and allowed us to visualise the details of the outflow pathways in the rabbit eyes. These results could be of great value in further evaluating the outcome of filtering surgeries in animal models.

Angiography↗
PubMed1982

Transcellular transport of fluorescein in hepatocyte monolayers: evidence for functional polarity of cells in culture.

The rat liver in vivo transfers bile salts, proteins, and dyes from blood into bile. It is the purpose of this communication to demonstrate the maintenance of this transcellular transport in cultured adult rat hepatocytes. Two minutes after adding fluorescein (20 microgram/ml) to the culture medium, maximal cellular fluorescence was observed through the fluorescence microscope. Subsequently, intercellular clefts showed a steadily increasing fluorescence with a maximum between 5 and 20 min, resulting in a brightly fluorescent network of intercellular gaps. The following observations are taken as evidence that these findings reflect cellular uptake and canalicular secretion of the dye. First, the same sequence of observations was made upon addition of fluorescein diacetate (a nonfluorescent precursor of fluorescein), proving that the compound had been taken up and metabolized in the cells to fluorescein before secretion into intercellular clefts. Second, preincubation of the monolayers with the cholestatic bile salt taurolithocholate (100 mumol/liter) suppressed almost completely intercellular but not cellular fluorescence. It is concluded that hepatocytes in culture show a functional polarity permitting the transcellular transport of substances bound for biliary secretion.

Animals↗
PubMed1997

Experimental and clinical study of detection of glioma at surgery using fluorescent imaging by a surgical microscope after fluorescein administration.

Total resection is the optimal treatment for malignant gliomas. However, an unexpected residual tumor mass is sometimes found on magnetic resonance imaging performed after an operation because of a macroscopically unclear margin of the tumor at surgery. This study was designed to evaluate the effectiveness of fluorescent imaging by a surgical microscope after fluorescein administration for the detection of gliomas at surgery. For this study, we produced two filters for the excitation and emission of fluorescein that can be easily fitted to and removed from a surgical microscope manually during the operation. For the experimental study, Wistar rat brains bearing C6 glioma were removed at appropriate intervals after intravenous administration of 10-20 mg kg-1 body weight of sodium fluorescein, and their surface and coronal section through the tumor were observed using a surgical microscope with the filters. In clinical cases, 1000 mg of sodium fluorescein was intravenously administered to five patients with glioma before tumor resection. In the experimental study, the C6 glioma itself and the edematous brain adjacent to the tumor (within 2-3 mm of the gross surface of the tumor) were well stained a brilliant yellowish green for a few hours. The normal brain was not stained. In clinical cases, the tumors were stained a brilliant yellowish green under fluorescent observation at surgery. The patients had no side effects. At all times the fluorescent observation could be quickly changed to ordinary observation by removing the filters from the surgical microscope. The tumor was also stained a faint yellow under ordinary nonfluorescent observation. Although this contributed to detection of the tumor, the fluorescent staining demarcated the tumor more clearly than nonfluorescent staining. These results suggest that this imaging technique by a surgical microscope with special filters at surgery may be practical and useful for detection of gliomas and warrants further clinical evaluations.

Animals↗
PubMed1987

Angiographic preoperative bowel segment localization using methylene blue, isosulfan blue, and fluorescein.

A canine model was used to determine the feasibility of preoperative, angiographic staining of bowel segments. Because methylene blue (MB) is contraindicated in patients with glucose-6-phosphate dehydrogenase deficiency or unstable hemoglobins, two alternate dyes, isosulfan blue (lymphazurin) and fluorescein were also studied. These three dyes were injected into 27 different mesenteric arterial branches in seven dogs following transfemoral catheterization. The duration of staining was evaluated over periods ranging from hours to 14 days, and pathologic sections were obtained to detect possible toxic effects. MB staining adequate for surgical resection persisted consistently up to 6 hours. Isosulfan blue and fluorescein produced adequate staining for only 30 to 60 minutes. Toxic mucosal changes were noted in four of 12 segments harvested within hours of the injection of MB and in none of the five MB injected segments harvested at four of 14 days. These changes varied greatly from focal regions of superficial necrosis to more generalized regions of transmucosal necrosis. All of the segments injected with isosulfan blue or fluorescein were free of histologic damage. It is concluded that the duration of staining produced by MB is sufficiently long to allow injection at angiography rather than at surgery. This allows decreased catheter time, and more importantly, precludes the possibility of interval catheter displacement. However, its clinical use should be viewed with some caution until its safety is established by further investigation. Isosulfan blue and fluorescein appear as alternate injectables, particularly for patients in whom MB is contraindicated. Because of their relatively short staining durations, injections with these agents should be made during surgery after angiographic catheter placement, although this method risks interval catheter dislodgement.

Angiography↗
PubMed1985

An evaluation of eight methods of using fluorescein to predict the viability of skin flaps in the pig.

The brightness of the fluorescence of fluorescein can be increased by the use of special (interference) filters that allow visualization of the dye when much smaller doses have been given. Standard random-pattern skin flaps were made in pigs and low (1 mg/kg) and high (500 mg) doses of fluorescein were given intravenously. The dye distribution in the flaps was evaluated visually, photographically, and with a dermofluorometer. All the methods proved reasonably accurate in predicting survival, although in an occasional flap, readings by all the methods were off by as much as 3.5 cm. The traditional Wood's lamp visualization proved to be as accurate as any of the newer methods. The inaccuracies of the fluorescein test are probably inherent in the method, since it only measures vascularity at the time the dye is given--and blood supply can change. The main advantage of low-dose fluorescein is that it allows repeated injections at reasonably close intervals. For serial observations, the dermofluorometer is unquestionably the best method.

Animals↗