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We retrospectively studied the value of iris fluorescein angiography in predicitng benignity or malignancy in 10 cases of ciliary body melanoma with iris extension. No correlation between fluorescein-angiographic appearance and histopathologic features was observed. Fluorescein angiography adds little to the management of ciliary body melanomas with iris extension.
We reviewed the clinical records, color photographs and fluorescein angiograms of 106 eyes of 53 patients (44 men and 9 women) with central serous chorioretinopathy who had been followed up for 1 year or more. Their ages ranged from 20 to 59 years. One eye in each patient was involved at the initial visit. According to fluorescein-angiographic findings, the lesions were divided into smokestack pattern (13 eyes), ink blot appearance (33 eyes) and minimally enlarging spot (7 eyes). Most patients complained of central scotoma, and most patients had moderately decreased visual acuities at the initial visit, which improved after recovery. In particular, all 7 eyes with a minimally enlarging spot had a visual acuity of 0.8 or more at the initial visit; of these 6 eyes had 1.0 or more several weeks later. On fluorescein angiography, 49 eyes demonstrated 1 leaking spot and 4 eyes showed 2 spots. Of 57 leaking spots, 10 were observed in the foveola, 31 were noted in the fovea and 16 were found in the para- or perifovea. Of 13 smokestack patterns, 5 were found in the inferonasal area. The ink blot appearance and minimally enlarging spots were frequently found in the superonasal and superotemporal areas.
The exact site of air leakage in a patient with primary spontaneous pneumothorax is difficult to determine and locate. In particular, the role of rupture of emphysema-like changes (blebs and bullae) versus that of enhanced porosity of lung parenchyma in the pathophysiology of primary spontaneous pneumothorax remains unclear. This is the first description of a patient with recurrent primary spontaneous pneumothorax in whom inhalation of aerosolized fluorescein followed by autofluorescence thoracoscopy allowed in vivo localization of various lung areas of extensive subpleural fluorescein accumulation which were not, or only partly, visibly abnormal during normal white light thoracoscopy. No air leak was present at the time of thoracoscopy. No emphysema-like changes were seen. Our findings suggest substantial areas of parenchymal abnormality that remain unnoticed by white light thoracoscopic inspection of the parenchymal surface. In this respect, fluorescein-enhanced autofluorescence thoracoscopy may become an exciting tool in the study of the pathophysiology of primary spontaneous pneumothorax, and could prove useful in clinical practice in determining the sites of surgical staple resection whenever this treatment modality is considered.
The esterases activity of normal and acute leukemic mouse lymphocytes and that of their homogenates was investigated using fluorescein diacetate (FDA) as a fluorogenic substrate. The activity proved to be the same for the two cell populations as well as for the homogenates prepared from them. In cell suspensions, having different osmolalities, the rate of FDA hydrolysis decreased significantly with the increasing osmolality only in the case of intact leukemic lymphocytes. changes in the membrane and cytoplasmic viscosity caused by increased or decreased environmental osmolality of cell suspensions occurred in the same direction and to the same extent for both normal and leukemic cells. Fluorescein, the fluorescent product of the hydrolysis, accumulates in leukemic lymphocytes, whereas it easily effluxes form normal lymphocytes. A flow microfluorimetry analysis of the cell population revealed that the fluorescein content of large leukemic lymphocytes was three times higher than that of small, normal ones. The observed differences specific for leukemic lymphocytes might be useful in detecting leukemic transformation in an early stage of acute lymphoid leukemia.
Expanding our previous finding of an adenosine-initiated transcription system, we now demonstrate that either the 5' site or the N6 site of adenosine nucleotides can be modified extensively without abolishing their ability to initiate transcription under the T7 phi2.5 promoter. Two series of amino derivatives of adenosine nucleotides were synthesized. Fluorescein and biotin groups were coupled to AMP derivatives through linkers of different sizes and hydrophobicities. Both fluorescein- and biotin-conjugated (at either the 5' or N6 site) adenosine nucleotides can act as efficient transcription initiators, producing fluorescein- and biotin-labeled RNA at the specific 5' end by a one-step transcription procedure, eliminating posttranscriptional modification. Furthermore, N6-modified adenosine derivative-initiated transcription synthesizes 5' end modified RNA with a free phosphate group, providing the possibility for further derivatization. The current finding makes easily available a variety of site-specifically functionalized RNA, which may be used in nucleic acid detection, RNA structural and functional investigation, and generation and isolation of novel functional RNA.
A general approach to detecting nucleic acid sequences in homogeneous media by means of steady-state fluorescence measurements is proposed. The methodology combines the use of a fluorescence-labeled single-strand DNA model probe, the complementary single-strand DNA target, and a DNA intercalator. The probe was fluorescein labeled to a spacer arm at the N4 position of the cytosine amino groups in polyribocytidylic acid (5'), poly(C), which acts as a model DNA probe. The complementary strand was polyriboinosinic acid (5'), poly(I), as a model of the target, and the energy transfer acceptor was an intercalator, either ethidium bromide or ethidium homodimer. In previous papers we have shown that the fluorescence intensity of the fluorescein label decreases when labeled poly(C) hybridizes with poly(I), and this fluorescence quenching can be used to detect DNA hybridization or renaturation in homogeneous media. In this paper we demonstrate that fluorescence resonance energy transfer (FRET) between fluorescein labeled to poly(C) and an intercalator agent takes place when single-stranded poly(C) hybridizes with poly(I), and we show how the fluorescence energy transfer further decreases the steady-state fluorescence intensity of the label, thus increasing the detection limit of the method. The main aim of this work was to develop a truly homogeneous detection system for specific nucleic acid hybridization in solution using steady-state fluorescence and FRET, but with the advantage of only having to label the probe with the energy donor since the energy acceptor is intercalated spontaneously. Moreover, the site label is not critical and can be labeled randomly in the DNA strand. Thus, the method is simpler than those published previously based on FRET. The experiments were carried out in both direct and competitive formats.
The anticalin FluA is an artificial lipocalin with novelspecificity for the fluorescein group, which was engineered from an insect bilin-binding protein by targeted random mutagenesis and selection. Based on the crystal structure of FluA, an attempt was made to improve the complementarity of its ligand pocket to fluorescein by rational protein design. Several side chains participating in sub-optimal interactions with the ligand were identified and replaced by residues that promised a better steric fit. As a result, the substitution of Ala45 by Ile and of Ser114 by Thr or Arg led to a tight affinity of ca. 1 nM, which is approximately 30-fold better than that of the parental anticalin. Similar to the original FluA, the improved version shows almost complete quenching of the bound ligand fluorescence. Interestingly, the quenching effect was significantly reduced when Trp129 was replaced by Tyr, thus supporting the previously postulated role of this residue, which closely packs against the bound ligand, for efficient electron transfer to the excited fluorescein. Circular dichroism spectra revealed that all variants investigated had retained the lipocalin fold. Corresponding thermal unfolding experiments confirmed similar folding stabilities, with melting temperatures ranging from 52.9 to 60.5 degrees C (i.e., for the high-affinity variant).
Fluorescein-PE is a fluorescence probe that is used as a membrane label or a sensor of surface associated processes. Fluorescein-PE fluorescence intensity depends not only on bulk pH, but also on the local electrostatic potential, which affects the local membrane interface proton concentration. The pH sensitivity and hydrophilic character of the fluorescein moiety was used to detect conformational changes at the lipid bilayer surface. When located in the dipalmitoylphosphatidylcholine (DPPC) bilayer, probe fluorescence depends on conformational changes that occur during phase transitions. Relative fluorescence intensity changes more at pretransition than at the main phase transition temperature, indicating that interface conformation affects the condition in the vicinity of the membrane. Local electrostatic potential depends on surface charge density, the local dielectric constant, salt concentration and water organisation. Initial increase in fluorescence intensity at temperatures preceding that of pretransition can be explained by the decreased value of the dielectric constant in the lipid polar headgroups region related in turn to decreased water organisation within the membrane interface. The abrupt decrease in fluorescence intensity at temperatures between 25 degrees C and 35 degrees C (DPPC pretransition) is likely to be caused by an increased value of the electrostatic potential, induced by an elevated value of the dielectric constant within the phosphate group region. Further increase in the fluorescence intensity at temperatures above that of the gel-liquid phase transition correlates with the calculated decreased surface electrostatic potential. Above the main phase transition temperature, fluorescence intensity increase at a salt concentration of 140 mM is larger than with 14 mM. This results from a sharp decline of the electrostatic potential induced by the phosphocholine dipole as a function of distance from the membrane surface.
We report the picosecond time-scale fluorescence dynamics of a dye-labeled DNA oligonucleotide or "aptamer" designed to bind specifically to Immunoglobulin E. Comparison of the photophysics of Texas Red (TR), fluorescein and 5'-carboxytetramethylrhodamine (TAMRA)-labeled aptamers reveals surprising differences with significant implications for measurements of oligonucleotide structure and dynamics. The fluorescence decay of the TR-aptamer is a simple single exponential with a weak temperature dependence. The fluorescence decay of the fluorescein-aptamer (fl-aptamer) is pH dependent and displays a complex temperature dependence with significant changes on melting of the aptamer tertiary structure. Despite its similarities to TR, TAMRA is strongly quenched when conjugated to the aptamer and displays complex fluorescence kinetics best described by a distributed rate model. Using the maximum entropy method, we have discovered two highly temperature-dependent fluorescence lifetimes for the TAMRA-aptamer. One of these lifetimes is similar to that of free TAMRA and displays the same temperature dependence. The other lifetime is quenched and displays a temperature dependence characteristic of a charge transfer reaction. These data set TR apart as an attractive alternative to TAMRA and fluorescein for studies such as fluorescence polarization and fluorescence resonance energy transfer, where environmental sensitivity of the probe is not desired.
PURPOSES: To evaluate the emergency resources of the clinics that perform fluorescein angiography in Pernambuco and, through a short multiple-choice questionnaire, to evaluate the ophthalmologists' performance to treat adverse reactions to contrast injection and to determine if the time after completing the specialization course in Ophthalmology influences in the results. METHODS: An interview with the eighteen ophthalmologists of the ten clinics where this examination is performed was made. The multiple-choice questionnaire had ten questions with five options each and only one correct answer. This questionnaire was about adverse reactions to fluorescein injection. RESULTS: One physician refused to participate and was excluded from the analysis. Among the others, fifteen physicians (88.2%) were certified specialists by the Brazilian Council of Ophthalmologists. All clinics had the basic structure for the initial care of adverse reactions to contrast injection. Half of these clinics (five) performed this examination with the presence of an anesthesiologist directly monitoring the patient and the other half had one on duty, if necessary. The number of correct answers varied between 3 and 8, with a mean of 5.2 +/- 1.6. Only four (23.6%) physicians gave more than seven correct answers. Specialists with five or less years of postgraduation in Ophthalmology had better results regarding the questionnaire (p<0.001). CONCLUSIONS: Despite appropriate material resources and support by anesthesiologist, the results of this study suggest that ophthalmologists who perform fluorescein angiography are not well-prepared to manage the possible adverse reactions, mainly doctors with more than five years after postgraduation in Ophthalmology.
The microsurgical anatomy of sellar regions in 20 cadavers with a 12 hours maximum post mortem was studied. Fluorescein was used as contrast for vascular anatomy examination. The use of fresh specimen and fluorescein had the objective of avoiding any distortion of normal anatomy. The drug was diluted with water or, in order to have more consistency, in gel for ultrasonography. Contrast injection was made into the internal carotid artery and photographs were taken with three different kinds of lighting: 1) only with Wood's lamp; 2) Wood's lamp and normal white light; 3) only with white light. Lighting only with Wood's lamp showed more contrast. After measurement, data often showed discrepancies with other authors' data. The photographic technique with fluorescein is simple and accessible; the drug showed high quality as a vascular contrast for neuroanatomic research.
Non-invasive tear film break-up time (NIBUT) was measured in nine normal subjects to investigate the effect of fluorescein instillation of tear film stability. It was found that fluorescein instillation reduced the tear film stability in the treated group, compared with the control group (P less than 0.05). It is, therefore likely that the tear film stability may be greater than had hitherto been suggested by the fluorescein method.
Video fluorescein angiography has been used to evaluate retinal circulatory parameters in diabetic and non-diabetic Sprague-Dawley rats. Video fluorescein angiograms were recorded from the retina using a modified retinal fundus camera following a 5 ul bolus injection of sodium fluorescein dye into the jugular vein. Retinal circulatory parameters were measured using computer assisted image analysis. These analyses were performed on 25 diabetic rats with 1 week duration of diabetes and 26 matched, non-diabetic, rats. There was a significant (p = .0001) increase in retinal Mean Circulation Time (MCT) in the diabetic group (1.83 +/- 0.40 s) compared to the control group (1.09 +/- 0.27 s). There were no significant differences in arterial or venous diameters comparing diabetic and control groups. In a separate paired experiment, measurements were made from the same animals both before and after one week duration of diabetes. A paired t-test analysis demonstrated significantly increased MCT times in the 6 diabetic animals (p = .001) while there was no significant differences detected in the 4 corresponding control animals. These results indicate that significant increases in retinal circulation times can be measured as early as 1 week after streptozotocin induced diabetes in this animal model.
Increasing interest in retinal research demands continuous improvement of experimental techniques and interpretation. Thus, the purpose of our research was to devise a new method for funduscopic photography and fluorescein angiography in the normal or diseased retina of the small laboratory animal that would produce results comparable in optical quality and field coverage to those obtained in human clinical practice. To enhance the view of the small eye, a 2.2 Volk Panretinal lens was held in apposition to the lens of a clinical fundus camera, the Topcon TRC 50FT, by means of a custom made metal sleeve. Albino mice, albino rats, and pigmented rats were photographed. Fluorescein angiography was performed on pigmented rats. Fluorescein was administered intravenously via the jugular vein at a dose of 5 mg/kg. Various speeds of film and flash settings were used depending on the light source and the pigmentation of the animal. Attachment of the 2.2 Panretinal lens to the clinical fundus camera allowed for more clearly defined fundus photographs of the small laboratory animal, as well as an enlarged field of observation over conventional techniques. Consequently, angiography fields and stages documented in the small laboratory animal approximated those obtained in human clinical practice. This technique facilitates the visualization of small fundi and it allows for a fuller documentation of experimental retinal models.
A method for evaluating the viability of mycobacterial cultures using fluorescein diacetate and ethidium bromide is described. Smears from 2-4 week old cultures of mycobacteria were stained with a 0.005% Dubos albumin broth dilution of fluorescein diacetate for thirty minutes at 37 C. The smears were counterstained with a 0.005% aqueous solution of ethidium bromide for ten minutes at room temperature and blotted dry. One drop of glycerol was placed on the smear and covered with a 22 X 40 mm coverslip. Stained smears were observed with fluorescence microscopy at 450 X. Viable organisms hydrolyzed the fluorescein diacetate and appeared yellow-green while dead organisms absorbed the ethidium bromide and appeared red-orange. Aliquots of material from which the slides were made were concurrently placed on Lowenstein-Jensen media and cultured for growth as a confirmation of viability. Investigation of 104 mycobacterial isolates using the described test showed that the viability of mycobacterial cultures can be determined by this method.
The clinical features and fluorescein angiographic findings of an 18-year-old man with PHPV are reported. The case is unusual in that the eye survived into adulthood. The lens resorbed spontaneously, thereby permitting study of the retrolental mass by fluorescein angiography. Fluorescein leakage from the persistent vessels was very slow but present, thus indicating a probable disruption of the endothelial tight junctions of the tunica vasculosa lentis.
BACKGROUND: Prophylaxis with oral Granisetron was assessed in patients undergoing fluorescein angiography (FAG) in order to evaluate its efficacy in reducing patients' discomfort due to nausea and vomiting (4% > 20%) and to ensure completion of the investigation. EXPERIMENTAL PLAN: Multicentre, randomised, double-blind, placebo-controlled study, with a follow-up visit performed 24 hrs after FAG. ENVIRONMENT: Five Anaesthesia/Resuscitation and Ophthalmology operational units participated in the study. PATIENTS: 120 patients (24 from each Centre) of both sexes and aged > 18 years, suffering from retinal disorders, were enrolled after informed consent. Pregnancy, lactation, anti-neoplastic treatment, ASA status IV/V were main exclusion criteria. All recruired patients completed the study. INTERVENTIONS: Ophthalmological evaluation was performed 30 days before and the same day as FAG. Anaesthesiological assessment on the same day as FAG, recording, BP, HR, ASA status, allergic diathesis, allergy to medications and contrast media, concomitant treatments. Oral Granisetrom 1 mg or placebo was given on an empty stomach 1 hr before the injection of 5 ml 20% fluorescein in 3 sec. A follow-up examination was done 24 hrs after FAG. SURVEY: In both groups nausea, retching and vomiting were seen after fluorescein injection. RESULTS: The study demonstrated that Granisetron is more active than placebo in controlling nausea (3 cases vs 4), and retching (2 vs 6). It was significantly more effective than placebo in the prevention of vomiting (0 vs 3). Efficacy was evaluated with chi 2, p < or = 0.05). CONCLUSIONS: The results suggest that Granisetron may be indicated as prophylaxis in patients undergoing FAG in order to prevent or control emesis, improve patients' comfort and ensure successful outcome of the investigation.
Fluorescein angiographic studies are reported on fifteen patients presenting with retinal opacification after blunt mechanical non-perforating injuries involving the eye. In patients who subsequently regained full visual function and in whom complete resolution of the retinal changes occurred, retinal fluorography carried out less than 24 hours after injury revealed no abnormal features. In cases with more prolonged visual disability, retinal fluorescein examinations may, however, reveal damage to the retinal pigment epithelium before this becomes apparent on routine ophthalmoscopy, or the existence of impaired perfusion through the retinal vessels. It is suggested, therefore, that retinal fluorescein angiography has a role in assessing the visual prognosis in patients presenting with traumatic retinal oedema.