RESUMO
The elucidation of the oral absorption of natural polysaccharides contributes to their further research and utilization. Herein, to explore the absorption of a pectin-type polysaccharide from Smilax china L. (SCLP), SCLP was respectively fluorescently labeled with fluorescein-5-thioicarbazide (FSCLP) and Cyanine7 amine (Cy7-SCLP) for in vitro and in vivo tracking. The near-infrared imaging demonstrated that Cy7-SCLP was absorbable in the small intestine and distributed in the liver and kidney after oral administration. Subsequently, in vitro intestinal epithelial tissue experiments showed that the jejunum was the dominant site of FSCLP transport. Further transport studies in the Caco-2 cell monolayer illustrated that FSCLP was delivered across the monolayer via transcellular transport by caveolae-mediated endocytosis and macropinocytosis together with paracellular transport by reversibly affecting tight junctions. In summary, this work presents the oral absorption characteristics and mechanisms of SCLP through the intestinal epithelium, which will facilitate the further development of SCLP and pectin polysaccharides.
Assuntos
Absorção Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Pectinas/farmacocinética , Polissacarídeos/farmacocinética , Smilax/química , Administração Oral , Animais , Células CACO-2 , Endocitose , Fluoresceína/administração & dosagem , Humanos , Mucosa Intestinal/efeitos dos fármacos , Rim/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Pectinas/administração & dosagem , Polissacarídeos/administração & dosagem , Ratos , Ratos Sprague-Dawley , Junções Íntimas , TranscitoseRESUMO
This work explored the feasibility of using biological polysaccharide to fabricate dissolvable microneedles (MNs) for the purpose of transdermal drug delivery and skin dendritic cell (DC) activation. Panax notoginseng polysaccharide (PNPS), a naturally derived immunoactive macromolecule, was used to fabricate dissolvable MNs. The prepared PNPS MNs showed a satisfactory mechanical strength and a skin penetration depth. By Franz diffusion cell assay, the PNPS MNs demonstrated a high transdermal delivery amount of model drugs. Furthermore, with the assistance of MNs, PNPS easily penetrated across the stratum corneum and target ear skin DCs, activating the maturation and migration of immunocytes by increasing the expressions of CD40, CD80, CD86, and MHC II of skin DCs. Consequently, the matured DCs migrated to the auricular draining lymph nodes and increased the proportions of CD4+ T and CD8+ T cells. Thus, PNPS might be a promising biomaterial for transdermal drug delivery, with adjuvant potential.
Assuntos
Células de Langerhans/efeitos dos fármacos , Agulhas , Panax notoginseng/química , Polissacarídeos/química , Administração Cutânea , Animais , Antígeno B7-1/metabolismo , Antígeno B7-2/metabolismo , Antígenos CD40/metabolismo , Força Compressiva , Doxorrubicina/administração & dosagem , Fluoresceína/administração & dosagem , Fluoruracila/administração & dosagem , Células de Langerhans/metabolismo , Masculino , Camundongos , Cadeias Pesadas de Miosina/metabolismo , Ratos Sprague-Dawley , Pele/citologia , Pele/efeitos dos fármacos , Pele/metabolismo , SolubilidadeRESUMO
AIM: To evaluate the effects of a proprietary formulated eye drop with Leptospermum spp honey versus a conventional lubricant eye drop on tear film properties in subjects with symptoms related to dry eye disease after 28 days of treatment. METHODS: Forty-six subjects with symptoms related to dry eye (Ocular Surface Disease Index (OSDI) score >12) were enrolled and randomly assigned to receive either the test formulated eye drop (Optimel by Melcare Biomedical Pty Ltd) or control eye drops (Alcon, USA) in this double-masked study. Inferior lipid layer thickness (LLT), tear film evaporation rate (TER), fluorescein tear film break-up time (TBUT), corneal staining and subjective symptoms (OSDI and visual analogue scales (VAS)) were measured before and after 28 days of instilling the eye drops. RESULTS: Forty-two subjects completed the study (21 subjects in each group). After 28 days of treatment, TER showed a significantly greater reduction with the formulated eye drop compared with the control (p=0.01). TBUT showed a slight but not statistically significant increase with the formulated eye drop compared with the control (p=0.06), and a significantly greater reduction (improvement) in OSDI scores was observed with the formulated eye drop compared with the control (p=0.01). No significant differences were found between the two groups for inferior LLT, corneal staining and any of the VAS scores. CONCLUSIONS: The formulated eye drops were effective in reducing tear film evaporation rate and were more effective for improving symptoms of dry eye compared with the control eye drops after 28 days of treatment. TRIAL REGISTRATION NUMBER: NCT03622619.
Assuntos
Apiterapia , Síndromes do Olho Seco/tratamento farmacológico , Mel , Leptospermum/química , Lubrificantes Oftálmicos/administração & dosagem , Lágrimas/fisiologia , Adolescente , Adulto , Método Duplo-Cego , Composição de Medicamentos , Síndromes do Olho Seco/fisiopatologia , Feminino , Fluoresceína/administração & dosagem , Corantes Fluorescentes/administração & dosagem , Humanos , Masculino , Preparações Farmacêuticas , Estudos Prospectivos , Coloração e Rotulagem , Inquéritos e Questionários , Lágrimas/química , Acuidade Visual/fisiologia , Adulto JovemRESUMO
PURPOSE: To compare the single application and two week treatment effects of device-applied (Eyepeace) and manually-applied eyelid massage techniques, as an adjunct to warm compress therapy, on ocular surface and tear film parameters. METHODS: Twenty participants (11 females, 9 males; mean age, 27⯱â¯11 years) with dry eye symptoms were recruited in a two week, investigator-masked, randomised, contralateral-eye trial. Following 10â¯min of warm compress therapy application (MGDRx EyeBag®) on both eyes, eyelid massage therapy was applied to one eye (randomised) by device, and to the fellow eye by manual eyelid massage, once daily for 14 days. Ocular surface and tear film measurements were conducted at baseline, and 15â¯min post-application by a clinician, then again after 14 days of self-administered daily treatment at home. RESULTS: Baseline clinical measurements did not differ between the treatment groups (all pâ¯>â¯0.05). Following two weeks of treatment, tear film lipid layer grade improved significantly with device massage (pâ¯=â¯0.008), and was marginally greater than manual massage by less than 1 grade (pâ¯=â¯0.03). Although immediate post-treatment improvements in tear film stability were observed in both groups (both pâ¯<â¯0.05), no significant long-term cumulative effects or inter-treatment differences in stability measures were detected (all pâ¯>â¯0.05). Visual acuity, tear meniscus height, conjunctival hyperaemia, ocular surface staining, and meibomian gland dropout did not change during the treatment period (all pâ¯>â¯0.05). CONCLUSIONS: Two weeks of treatment with the eyelid massage device, as an adjunct to warm compress therapy, effected marginally greater improvements in tear film lipid layer thickness than the conventional manual technique, which were statistically but not clinically significant. Future parallel group trials with longer treatment periods and a greater range of disease severity are required.
Assuntos
Síndromes do Olho Seco/terapia , Pálpebras/fisiologia , Massagem/instrumentação , Disfunção da Glândula Tarsal/terapia , Adulto , Corantes/administração & dosagem , Método Duplo-Cego , Síndromes do Olho Seco/fisiopatologia , Pálpebras/efeitos dos fármacos , Feminino , Fluoresceína/administração & dosagem , Corantes Fluorescentes/administração & dosagem , Humanos , Corantes Verde de Lissamina/administração & dosagem , Masculino , Disfunção da Glândula Tarsal/fisiopatologia , Estudos Prospectivos , Inquéritos e Questionários , Lágrimas/fisiologia , Adulto JovemRESUMO
PURPOSE: To test the safety and efficacy of high frequency electrotherapy (ET) on the clinical signs and symptoms of patients affected by dry eye and meibomian gland dysfunction (MGD). METHODS: Twenty-five patients affected by MGD were enrolled. Quantum Molecular Resonance ET was administered by means of the Rexon-Eye device 4 times, once per week for 4 weeks. Patients were reexamined 1 month after the last treatment. The primary endpoint was reduction in corneal fluorescein staining. Additional endpoints were tear break-up time, Ocular Surface Disease Index score, meibomian gland secretion score, and the number of expressible meibomian glands. Safety endpoints were Logarithm of the Minimum Angle of Resolution (LogMar) best spectacle-corrected visual acuity and intraocular pressure. RESULTS: Corneal fluorescein staining improved by 62.5% (P < 0.0001), tear breakup time increased by 30.9% (P < 0.0001), and the Ocular Surface Disease Index score improved by 37% (P < 0.001). The meibum quality and the number of expressible meibomian glands also increased (35.7% and 12%, P < 0.001 and P < 0.0001, respectively). Schirmer test scores increased after treatment by 16.5% (P = 0.01). No adverse events were observed. CONCLUSIONS: Quantum Molecular Resonance ET appears to be safe and significantly reduces symptoms and signs associated with MGD. It may have a relevant role in the treatment of evaporative dry eye disease.
Assuntos
Córnea/patologia , Terapia por Estimulação Elétrica/instrumentação , Disfunção da Glândula Tarsal/terapia , Glândulas Tarsais/metabolismo , Lágrimas/metabolismo , Córnea/metabolismo , Desenho de Equipamento , Feminino , Fluoresceína/administração & dosagem , Corantes Fluorescentes/farmacologia , Seguimentos , Humanos , Masculino , Disfunção da Glândula Tarsal/diagnóstico , Disfunção da Glândula Tarsal/metabolismo , Pessoa de Meia-Idade , Soluções Oftálmicas , Estudos Prospectivos , Resultado do TratamentoRESUMO
PURPOSE: The blood-retina barrier (BRB) is a biological barrier consisting of tightly interconnected endothelial cells inside the retinal vascular network that protects the neural tissue from harmful pathogens and neurotoxic molecules circulating in the bloodstream. Unfortunately, with regard to retinoblastoma, this barrier also prevents systemically administered therapeutics reaching the retinal tissue. In this study we introduce a novel technique to locally and transiently increase BRB permeability for drug delivery using hyperthermia of magnetic nanoparticles (MNPs). MATERIALS AND METHODS: An alternating current (AC) magnetic field was used to induce hyperthermia of locally injected MNPs in the left ophthalmic artery of a rat model. To improve adherence on the surface of the endothelium, commercially available MNPs coated with human transferrin glycoproteins were used. After hyperthermia we assessed the extravasation of systemically injected sodium fluorescein (NaF) as well as Evans blue dye (EBD) into the retinal tissue. RESULTS: Spectrofluorometry and fluorescent microscopy image analysis show a significant increase of dye penetration in the retina where hyperthermia of MNPs was applied. CONCLUSIONS: Our proposed new technique can allow both small and large dye molecules to cross the BRB. While the results are preliminary and thorough evaluation of the retinal tissue following hyperthermia is necessary, this technique has the potential to be an effective mean for the treatment of various diseases such as retinoblastoma.
Assuntos
Barreira Hematorretiniana/metabolismo , Corantes/administração & dosagem , Azul Evans/administração & dosagem , Fluoresceína/administração & dosagem , Hipertermia Induzida , Nanopartículas de Magnetita/administração & dosagem , Animais , Campos Magnéticos , Masculino , RatosRESUMO
BACKGROUND: Pulmonary drug delivery is characterized by short onset times of the effects and an increased therapeutic ratio compared to oral drug delivery. This delivery route can be used for local as well as for systemic absorption applying drugs as single substance or as a fixed dose combination. Drugs can be delivered as nebulized aerosols or as dry powders. A screening system able to mimic delivery by the different devices might help to assess the drug effect in the different formulations and to identify potential interference between drugs in fixed dose combinations. The present study evaluates manual devices used in animal studies for their suitability for cellular studies. METHODS: Calu-3 cells were cultured submersed and in air-liquid interface culture and characterized regarding mucus production and transepithelial electrical resistance. The influence of pore size and material of the transwell membranes and of the duration of air-liquid interface culture was assessed. Compounds were applied in solution and as aerosols generated by MicroSprayer IA-1C Aerosolizer or by DP-4 Dry Powder Insufflator using fluorescein and rhodamine 123 as model compounds. Budesonide and formoterol, singly and in combination, served as examples for drugs relevant in pulmonary delivery. RESULTS AND CONCLUSIONS: Membrane material and duration of air-liquid interface culture had no marked effect on mucus production and tightness of the cell monolayer. Co-application of budesonide and formoterol, applied in solution or as aerosol, increased permeation of formoterol across cells in air-liquid interface culture. Problems with the DP-4 Dry Powder Insufflator included compound-specific delivery rates and influence on the tightness of the cell monolayer. These problems were not encountered with the MicroSprayer IA-1C Aerosolizer. The combination of Calu-3 cells and manual aerosol generation devices appears suitable to identify interactions of drugs in fixed drug combination products on permeation.
Assuntos
Broncodilatadores/administração & dosagem , Broncodilatadores/farmacocinética , Avaliação Pré-Clínica de Medicamentos/instrumentação , Avaliação Pré-Clínica de Medicamentos/métodos , Administração por Inalação , Aerossóis/administração & dosagem , Aerossóis/farmacocinética , Budesonida/administração & dosagem , Budesonida/farmacocinética , Combinação Budesonida e Fumarato de Formoterol/administração & dosagem , Combinação Budesonida e Fumarato de Formoterol/farmacocinética , Membrana Celular/efeitos dos fármacos , Células Cultivadas , Combinação de Medicamentos , Células Epiteliais/efeitos dos fármacos , Desenho de Equipamento , Fluoresceína/administração & dosagem , Fluoresceína/farmacocinética , Fumarato de Formoterol/administração & dosagem , Fumarato de Formoterol/farmacocinética , Humanos , Nebulizadores e Vaporizadores , Rodamina 123/administração & dosagem , Rodamina 123/farmacocinéticaRESUMO
Certain species of urodeles and teleost fish can regenerate their tissues. Zebrafish have become a widely used model to study the spontaneous regeneration of adult tissues, such as the heart, retina, spinal cord, optic nerve, sensory hair cells, and fins. The zebrafish fin is a relatively simple appendage that is easily manipulated to study multiple stages in epimorphic regeneration. Classically, fin regeneration was characterized by three distinct stages: wound healing, blastema formation, and fin outgrowth. After amputating part of the fin, the surrounding epithelium proliferates and migrates over the wound. At 33 °C, this process occurs within six hours post-amputation (hpa, Figure 1B). Next, underlying cells from different lineages (ex. bone, blood, glia, fibroblast) re-enter the cell cycle to form a proliferative blastema, while the overlying epidermis continues to proliferate (Figure 1D). Outgrowth occurs as cells proximal to the blastema re-differentiate into their respective lineages to form new tissue (Figure 1E). Depending on the level of the amputation, full regeneration is completed in a week to a month. The expression of a large number of gene families, including wnt, hox, fgf, msx, retinoic acid, shh, notch, bmp, and activin-betaA genes, is up-regulated during specific stages of fin regeneration. However, the roles of these genes and their encoded proteins during regeneration have been difficult to assess, unless a specific inhibitor for the protein exists, a temperature-sensitive mutant exists or a transgenic animal (either overexpressing the wild-type protein or a dominant-negative protein) was generated. We developed a reverse genetic technique to quickly and easily test the function of any gene during fin regeneration. Morpholino oligonucleotides are widely used to study loss of specific proteins during zebrafish, Xenopus, chick, and mouse development. Morpholinos basepair with a complementary RNA sequence to either block pre-mRNA splicing or mRNA translation. We describe a method to efficiently introduce fluorescein-tagged antisense morpholinos into regenerating zebrafish fins to knockdown expression of the target protein. The morpholino is micro-injected into each blastema of the regenerating zebrafish tail fin and electroporated into the surrounding cells. Fluorescein provides the charge to electroporate the morpholino and to visualize the morpholino in the fin tissue. This protocol permits conditional protein knockdown to examine the role of specific proteins during regenerative fin outgrowth. In the Discussion, we describe how this approach can be adapted to study the role of specific proteins during wound healing or blastema formation, as well as a potential marker of cell migration during blastema formation.
Assuntos
Eletroporação/métodos , Morfolinos/administração & dosagem , Oligonucleotídeos Antissenso/administração & dosagem , Regeneração/fisiologia , Peixe-Zebra/fisiologia , Nadadeiras de Animais/fisiologia , Animais , Fluoresceína/administração & dosagem , Corantes Fluorescentes/administração & dosagem , Regeneração/genética , Cauda/fisiologia , Peixe-Zebra/genéticaRESUMO
The vomeronasal system is segregated from the epithelium to the bulb. Two classes of receptor neurons are apically and basally placed in the vomeronasal epithelium, express Gi2alpha and Goalpha proteins and V1R and V2R receptors and project to the anterior and posterior portions of the accessory olfactory bulb, respectively. Apart from common vomeronasal recipient structures in the amygdala, only the anterior accessory olfactory bulb projects to the bed nucleus of the stria terminalis and only the posterior accessory olfactory bulb projects to the dorsal anterior amygdala. The efferent projections from these two amygdaloid structures to the hypothalamus were investigated. These two vomeronasal subsystems mediated by V1R and V2R receptors were partially segregated, not only in amygdala, but also in the hypothalamus.
Assuntos
Vias Aferentes/fisiologia , Vias Eferentes/fisiologia , Hipotálamo/fisiologia , Receptores de Vasopressinas/metabolismo , Órgão Vomeronasal/fisiologia , Vias Aferentes/anatomia & histologia , Vias Aferentes/metabolismo , Aminas/administração & dosagem , Aminas/farmacocinética , Tonsila do Cerebelo/anatomia & histologia , Tonsila do Cerebelo/metabolismo , Tonsila do Cerebelo/fisiologia , Animais , Dextranos/administração & dosagem , Dextranos/farmacocinética , Vias Eferentes/anatomia & histologia , Vias Eferentes/metabolismo , Feminino , Fluoresceína/administração & dosagem , Fluoresceína/farmacocinética , Corantes Fluorescentes/administração & dosagem , Corantes Fluorescentes/farmacocinética , Hipotálamo/anatomia & histologia , Hipotálamo/metabolismo , Masculino , Microinjeções , Bulbo Olfatório/anatomia & histologia , Bulbo Olfatório/metabolismo , Bulbo Olfatório/fisiologia , Ratos , Ratos Sprague-Dawley , Rodaminas/administração & dosagem , Rodaminas/farmacocinética , Núcleos Septais/anatomia & histologia , Núcleos Septais/metabolismo , Núcleos Septais/fisiologia , Órgão Vomeronasal/anatomia & histologia , Órgão Vomeronasal/metabolismoRESUMO
OBJECT: The authors performed fluorescein cerebral angiography in patients after aneurysm clip placement to confirm the patency of the parent artery, perforating artery, and other arteries around the aneurysm. METHODS: Twenty-three patients who underwent aneurysm surgery were studied. Aneurysms were located in the internal carotid artery in 12 patients, middle cerebral artery in six, anterior cerebral artery in three, basilar artery bifurcation in one, and junction of the vertebral artery (VA) and posterior inferior cerebellar artery in one. After aneurysm clip placement, the target arteries were illuminated using a beam from a blue light-emitting diode atop a 7-mm diameter pencil-type probe. In all patients, after intravenous administration of 5 ml of 10% fluorescein sodium, fluorescence in the vessels was clearly observed through a microscope and recorded on videotape. RESULTS: The excellent image quality and spatial resolution of the fluorescein angiography procedure facilitated intraoperative real-time assessment of the patency of the perforating arteries and branches near the aneurysm, including: 12 posterior communicating arteries; 12 anterior choroidal arteries; four lenticulostriate arteries; three recurrent arteries of Heubner; three hypothalamic arteries; one ophthalmic artery; one perforating artery arising from the VA; and one posterior thalamoperforating artery. All 23 patients experienced an uneventful postoperative course without clinical symptoms of perforating artery occlusion. CONCLUSIONS: Because the fluorescein angiography procedure described here allows intraoperative confirmation of the patency of perforating arteries located deep inside the surgical field, it can be practically used for preventing unexpected cerebral infarction during aneurysm surgery.
Assuntos
Aneurisma Roto , Encéfalo/irrigação sanguínea , Angiografia Cerebral/métodos , Fluoresceína , Aneurisma Intracraniano , Dissecação da Artéria Vertebral , Aneurisma Roto/diagnóstico por imagem , Aneurisma Roto/fisiopatologia , Aneurisma Roto/cirurgia , Velocidade do Fluxo Sanguíneo , Circulação Cerebrovascular/fisiologia , Meios de Contraste , Desenho de Equipamento , Fluoresceína/administração & dosagem , Humanos , Hipotálamo/irrigação sanguínea , Injeções Intravenosas , Aneurisma Intracraniano/diagnóstico por imagem , Aneurisma Intracraniano/fisiopatologia , Aneurisma Intracraniano/cirurgia , Cuidados Intraoperatórios , Procedimentos Neurocirúrgicos/instrumentação , Artéria Oftálmica/fisiopatologia , Artéria Oftálmica/cirurgia , Tálamo/irrigação sanguínea , Tálamo/fisiopatologia , Tálamo/cirurgia , Dissecação da Artéria Vertebral/diagnóstico por imagem , Dissecação da Artéria Vertebral/fisiopatologia , Dissecação da Artéria Vertebral/cirurgiaRESUMO
Presented in the paper are data of a comparative analysis of efficiency of different methods of administration of drugs in neuritis and partial atrophy of the optic nerve. New techniques of application and fixation of irrigation systems in the retrobulbar and Tenon's space are described. Experimental and clinical data proving advantages of the new method of administration of drugs by an automatic pulse doser in the treatment of inflammatory diseases of the optic nerve are represented. The use of such intensive intermittent technique of administration of drugs in Tenon's space performed at the preliminary stage before electrostimulation of the optic nerve made the procedure by far more effective and ensured better treatment results.
Assuntos
Antibacterianos/administração & dosagem , Diuréticos/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Atrofia Óptica/tratamento farmacológico , Neurite Óptica/tratamento farmacológico , Inibidores de Proteases/administração & dosagem , Animais , Criança , Pré-Escolar , Meios de Contraste/administração & dosagem , Modelos Animais de Doenças , Quimioterapia Combinada , Feminino , Fluoresceína/administração & dosagem , Humanos , Lactente , Bombas de Infusão Implantáveis , Masculino , Atrofia Óptica/patologia , Atrofia Óptica/fisiopatologia , Disco Óptico/efeitos dos fármacos , Disco Óptico/patologia , Neurite Óptica/patologia , Neurite Óptica/fisiopatologia , Órbita , Coelhos , Irrigação Terapêutica/instrumentação , Acuidade Visual/fisiologiaRESUMO
The purpose of this work is to validate a novel ocular microdialysis sampling technique in rabbits with permanently implanted vitreous probes. This objective is achieved by studying the vitreous pharmacokinetics of fluorescein following systemic and intravitreal administration. The rabbits were divided into two groups (groups I and II) based on whether or not they were allowed a recovery period following surgical implantation of probes. The integrity of the blood-retinal barrier was determined by the vitreal protein concentrations and the fluorescein permeability index. Vitreal protein concentrations returned to baseline 48 h after probe implantation and therefore experiments were conducted 72 h post-implantation of probes in rabbits where recovery period was allowed. The permeability indices for fluorescein after systemic administration in group I (without recovery period) and group II (with recovery period) indicated that the integrity of the blood-retinal barrier was maintained and were found out to be 0.55 +/- 0.27 and 0.71 +/- 0.38%, respectively, for the vitreous chamber. Following microdialysis probe implantation in the group II rabbits, the blood-retinal barrier integrity was not compromised. A novel microdialysis technique in rabbits with permanently implanted probes for studying the pharmacokinetics of posterior segment has been developed and characterized.
Assuntos
Cateteres de Demora/tendências , Fluoresceína/farmacocinética , Microdiálise/métodos , Corpo Vítreo/efeitos dos fármacos , Animais , Barreira Hematorretiniana/efeitos dos fármacos , Barreira Hematorretiniana/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Olho/efeitos dos fármacos , Olho/metabolismo , Fluoresceína/administração & dosagem , Meia-Vida , Injeções/métodos , Masculino , Modelos Animais , Cuidados Pós-Operatórios/métodos , Coelhos , Fatores de Tempo , Corpo Vítreo/química , Corpo Vítreo/metabolismoRESUMO
BACKGROUND: The tear function index (TFI) has been shown to be of value in the diagnosis of patients suffering from Sjögren's syndrome. It is dependent, however, on introducing into the conjunctival fornix the correct concentration of fluorescein in at least one and a half times the normal tear volume. The stimulus and effect of this added volume on the tear dynamics is likely to vary between individuals. These factors, together with the method of performing the test, limit its general applicability. AIM: To devise a method of performing the TFI with less variability and more general applicability. To present a theoretical and in vitro assessment of the dynamics of the TFI. METHOD: The study was divided into three parts. The first part was to compare the results obtained using a prepared strip containing 1.3 microl of 0.5% fluorescein with the introduction of the same amount of fluorescein as a drop. The second part was to compare the results obtained with prepared strips with the standard method of performing the TFI, both with and without topical anaesthetic. The third part was an in vitro study of the rate of flow of graded volumes on a filter paper strip. 42 subjects with a diagnosis of Sjögren's syndrome according to the European criteria and 126 without Sjögren's syndrome were included. RESULTS: There was no significant difference between the results obtained with a prepared strip and the introduction of 1.3 microl into the eye before performing the Schirmer's test and TFI (0.1