Beta-cell Antibody Research Articles

THU354 Enfortumab Vedotin Induced Refractory Hyperglycemia

Abstract Disclosure: M. Hussein: None. Introduction: Immunotherapy has transformed the treatment landscape of cancer therapy and has created promising hope for cancer patients. While immunotherapy has dramatic efficacy it also has unique toxicity profile. A spectrum of adverse events known as endocrinopathies have been reported with immune checkpoint inhibitors. One of the newer medications is Enfortumab Vedotin, an antibody-drug conjugate. This drug is approved for use in advanced urothelial carcinoma. Case: We present a 68-year-old male with advanced urothelial carcinoma who was started on Enfortumab vedotin after he progressed on a different chemotherapy/immunotherapy. His medical history was significant for prediabetes and morbid obesity.2 weeks after the first cycle of treatment he presented to emergency room with worsening dyspnea and fatigue. Vital signs: Temp: 37.3, HR: 90, BP: 122/65, RR 18, SPO2 95% on 3L O2 through nasal canula. BMI 44 kg/m. Biochemical workup revealed elevated Glucose level of 310 and anion gap of 12. He was started on subcutaneous insulin. He continued to be uncontrolled and was transferred to the ICU and switched to intravenous insulin due to hyperglycemia. His total daily insulin dose was close to 900 units to achieve blood glucose <180. It took 48-72 hours of Intravenous insulin use to get controlled blood glucose. Hgba1c was 7.3%. Insulin antibody, GAD65 and Islet cell antibodies were negative, C peptide 38 for a glucose of 428. Unfortunately, the patient developed multiorgan failure, and passed away on day 7 of the admission. Discussion: The mechanism of this drug inducing hyperglycemia is not well understood as of yet. But the presumed theory here is the patient developed severe acute insulin resistance that led to this refractory hyperglycemia, as evident by the high c peptide level. Absence of beta cell antibodies and elevated c peptide go against an autoimmune destructive beta cell pathway. The other theory is that there is a malfunction in the signaling pathway at the cellular level that led to hyperglycemia. Risk of developing hyperglycemia as a side effect to this medication increase with patient’s personal risk factors for developing diabetes mellitus (prediabetes and obesity). In phase 3 trial of Enfortumab vedotinEV301, treatment related hyperglycemia was reported in 6.4 % of patients. Hyperglycemia occurred more frequently in patients with hyperglycemia at baseline or with a body-mass index of 30 or higher. Conclusion: We are in need for further studies to understand the mechanism of refractory hyperglycemia induced by this drug. Patients on this type of therapy should be monitored very closely for developing a new onset of diabetes mellitus or worsening of established diabetes mellitus. Presentation: Thursday, June 15, 2023

Disease heterogeneity of adult diabetes based on routine clinical variables at diagnosis: Results from the German/Austrian Diabetes Follow-up Registry.

To cluster adults with diabetes using variables from real-world clinical care at manifestation. We applied hierarchical clustering using Ward's method to 56 869 adults documented in the prospective Diabetes Follow-up Registry (DPV). Clustering variables included age, sex, body mass index (BMI), HbA1c, diabetic ketoacidosis (DKA), components of the metabolic syndrome (hypertension/dyslipidaemia/hyperuricaemia) and beta-cell antibody status. Time until use of oral antidiabetic drugs (OADs), use of insulin, chronic kidney disease (CKD), cardiovascular disease (CVD), retinopathy or neuropathy were assessed using Kaplan-Meier analysis and Cox regression models. We identified eight clusters: four clusters comprised early diabetes onset (median age 40-50 years) but differed with regard to BMI, HbA1c, DKA and antibody positivity. Two clusters included adults with diabetes onset aged in their early 60s who met target HbA1c, but differed in BMI and sex distribution. Two clusters were characterized by late diabetes onset (median age 69 and 77 years) and comparatively low BMI, but differences in HbA1c. Earlier insulin use was observed in adults with high HbA1c, and earlier OAD use was observed in those with high BMI. Time until CKD or CVD was shorter in those with late onset, whereas retinopathy occurred earlier in adults with late onset and high HbA1c, and in adults with early onset, but high HbA1c and high percentage of antibody positivity. Adult diabetes is heterogeneous beyond classical type 1/type 2 diabetes, based on easily available variables in clinical practice using an automated clustering algorithm that allows both continuous and binary variables.

Clinical and laboratory clues of maturity-onset diabetes of the young and determination of association with molecular diagnosis.

Maturity-onset diabetes of the young (MODY) is often misdiagnosed as other types of diabetes because it is overlooked due to atypical clinical presentations. This study aims to reveal the clinical and laboratory clues and examine their compatibility with MODY genotypes. Participants consisted of 230 children with atypical presentations for type1(T1DM) and type2 diabetes mellitus (T2DM). MODY-causing mutations were screened in the following genes:GCK-HNF1A-HNF4A-HNF1B-PDX1-NEUROD1-KLF11-CEL-PAX4-INS-BLK. Clinical and laboratory features were compared between children with MODY and children without MODY. The most common reasons for MODY screening were as follows (n/%):low daily dose of insulin (DDI) requirement (122/53%), absence of beta-cell antibodies(58/25.3%), coincidental hyperglycemia(26/11.3%), family history of diabetes (12/5.2%), hypoglycemia/hyperglycemia episodes(7/3%), hyperglycemia related to steroids(3/1.4%) and renal glycosuria(2/0.8%). The markers with the most likelihood to distinguish MODY from T1DM were determined as follows: measurable C-peptide in follow-up, family history of early-onset diabetes and low DDI requirement (odds ratio:12.55, 5.53 and 3.43, respectively). The distribution of the most common causative genes in children with MODY(n = 24) is as follows (n/%):GCK(15/62.5%), HNF4A(7/29.1%), HNF1A(1/9.2%) and PDX1(1/9.2%).All children(n = 12) with GCK-MODY(MODY2) were screened for low DDI requirement, while beta-cell negativity was more common in HNF4A-MODY(MODY1). The study shows that measurable C-peptide in follow-up, family history of early-onset diabetes, and low DDI are still remarkable clues to predict MODY in children with misdiagnosed T1DM. In addition, the most common mutations were found in the GCK and HNF4A genes. Among children misdiagnosed with T1DM, a low DDI requirement was found more frequently in MODY2, whereas beta-cell antibody negativity was more common in MODY1.

Effect of oral insulin on prevention of diabetes in relatives of patients with T1DM: a randomized clinical trial

Insulin is one of the autoantigens involved in the pathogenesis of T1DM. Therefore several trials with insulin given orally, nasal or subcutaneous have been conducted to prevent the development of T1DM in genetically high risk populations. This trial design built on the results of DPT-1 and included further investigations related to first phase insulin release. A dose of 7.5 mg/d did not prevent T1DM in the high risk population of autoantibody positive T1DM relatives. Possibly ongoing trials with higher insulin doses might show better effects on beta cell autoimmunity and T1DM incidence. Knowledge of pre-diabetes staging has informed several prevention trials and epidemiological studies in high risk populations. Individuals with two or more beta cell antibodies are now classified as Stage 1. Without intervention, most of these will develop abnormal glucose tolerance (Stage 2) and then clinically manifest T1DM (Stage 3). The overall rate of progression from Stage 1 to 3 is ~9.5% per year.

Differential diagnostic utilities of combined testing for islet cell antibody, glutamic acid decarboxylase antibody, and tyrosine phosphatase antibody

Background. Beta-cell antibody tests are used for the differential diagnosis of diabetes mellitus. They permit to discriminate between the type 1 diabetes (T1D) and non-autoimmune diabetes types. To choose an appropriate test for ruling in or ruling out the T1D a physician needs to know how conclusive test results are. The most powerful estimate of test conclusiveness is its likelihood ratio (LHR). The higher LHR of a positive result (LHR+), the more posttest probability of T1D; the lower LHR of a negative result (LHR), the less posttest probability of T1D.
 Aims. To compare conclusiveness of single and combined tests for antibodies to islet cells (ICA), glutamate decarboxylase (GADA), and tyrosine phosphatase IA-2 (IA-2A), and to evaluate posttest probabilities of T1D at various pretest probabilities.
 Methods. All antibodies were tested in parallel in 169 children and adolescents with a new-onset T1D, and in 169 persons without this disease. ICA, GADA, and IA-2A were determined by indirect immunofluorescence, radioimmune assay, and ELISA, respectively. LHR+ and LHR were calculated with the MedCalc Statistical Software. Posttest T1D probabilities were calculated from Bayes theorem-based equation.
 Results. Among single tests, an ICA test had the greatest LHR+ and the smallest LHR, and consequently was the most reliable either for ruling in or ruling out the T1D. Among test combinations, an ICAGADA combination had the greatest LHR+ and was the most suitable for T1D confirmation. The triple combination ICAGADAIA-2A had the smallest LHR and was the most suitable for T1D exclusion.
 Conclusions. In the differential diagnosis of diabetes, the most appropriate test for ruling in the T1D is the double combination ICAGADA. With both antibodies positive, this combination provides the highest posttest T1D probabilities at any pretest probability. The most appropriate test for ruling out the T1D is the triple combination ICAGADAIA-2A. With all three antibodies negative, this combination provides the lowest posttest T1D probabilities.

Maturity onset diabetes of the young due to HNF1A variants in Croatia.

IntroductionMaturity onset diabetes of the young due to HNF1A mutations (HNF1A-MODY) is the most frequent form of monogenic diabetes in adults. It is often misdiagnosed as type 1 or type 2 diabetes, but establishing genetic diagnosis is important, as treatment differs from the common types of diabetes. HNF1A-MODY has not been investigated in Croatia before due to limited access to genetic testing. In this study we aimed to describe the characteristics of young adults diagnosed with diabetes before the age of 45 years, who have rare HNF1A allele variants, and estimate the prevalence of HNF1A-MODY in Croatia.Materials and methodsWe recruited 477 C-peptide positive and beta cell antibody negative subjects through the Croatian Diabetes Registry. HNF1A was sequenced for all participants and systematic assessment of the variants found was performed. The prevalence of HNF1A-MODY was calculated in the study group and results extrapolated to estimate the proportion of diabetic individuals with HNF1A-MODY in Croatia and the population prevalence.ResultsOur study identified 13 individuals harbouring rare HNF1A allelic variants. After systematic assessment, 8 were assigned a diagnosis of HNF1A-MODY. Two individuals were able to discontinue insulin treatment following the diagnosis. We estimated that HNF1A-MODY in Croatia has a prevalence of 66 (95% CI 61 - 72) cases per million.ConclusionsThe estimated prevalence of HNF1A-MODY in Croatia is similar to that reported in other European countries. Finding cases lead to important treatment changes for patients. This strongly supports the introduction of diagnostic genetic testing for monogenic diabetes in Croatia.

The prognostic significance of glutamic acid decarboxylase antibodies in patients with chronic pancreatitis undergoing total pancreatectomy with islet autotransplantation

AimIslet autotransplantation (IAT) is considered a ‘non-immune’ model of islet transplant, with no risk for autoimmune-mediated beta cell loss, but we have previously observed de novo type 1 diabetes in one total pancreatectomy with islet autotransplantation (TPIAT) recipient. We aimed to investigate the clinical significance of glutamic acid decarboxylase antibodies (GADA), as a sensitive marker for autoimmune diabetes mellitus (DM), in patients with chronic pancreatitis undergoing TPIAT. MethodsWe identified 9 patients undergoing TPIAT with elevated GADA pre-TPIAT (8 non-diabetic and 1 with C-peptide positive DM), otherwise demographically similar to GADA negative TPIAT recipients (n=341). Metabolic and clinical measures related to islet cell function were recorded both before and after TPIAT. ResultsNone of the 9 TPIAT patients achieved insulin independence after surgery, vs. 33% of GADA negative patients (n=318 with 1-yr follow-up). The two patients with the highest titters of GADA (>250 IU/mL) both experienced islet graft failure, despite normoglycaemia pre-TPIAT and high islet mass transplanted (5276 and 9378 IEQ per kg), with elevated HbA1c levels post-TPIAT (8.3%, 9.6%). The remaining 7 seven were insulin dependent with partial graft function and HbA1c levels <7%. ConclusionInsulin dependence was more frequent in 9 patients with elevated GADA prior to TPIAT than in GADA negative TPIAT recipients, with graft failure in 2 cases. We speculate that beta-cell autoimmunity may occur in a small subset of TPIAT recipients and that beta cell antibody testing prior to TPIAT may be warranted to identify individuals at higher risk for insulin dependence.

Protective effect and potential mechanisms of propolis on streptozotocin-induced diabetic rats

Abstract Objectives The objective of this study was to determine the protective effects of propolis supplementation against the toxic effect of streptozotocin (STZ) on pancreatic beta-cells and to identify the possible mechanism of actions underlying this effect. Methods Forty-five rats were randomly divided into three groups, with each group containing 15 rats. Group I consisted of normal rats fed a normal chow diet. Group II included diabetic rats induced with STZ. Group III consisted of rats treated with 0.3 g/Kg/day propolis for 2 weeks before the induction of diabetes by STZ. At the end of experimental period, blood samples were collected for the measurement of fasting blood glucose (FBG) and lipid peroxidation activity. Pancreatic tissues from the dissected rats were processed for morphological and immunohistochemical studies. Results The findings of this work showed that the treatment of rats with propolis before the induction of diabetes mellitus is associated with significantly decreased FBG levels compared to the diabetic rats. The lipid peroxidation products were significantly increased in group II compared to group I. However, these products were significantly decreased in group III compared to group II. The histopathological and immunohistochemical analysis of the pancreatic tissue showed an increase in the size of the islets of Langerhans and the enhanced staining of beta-cells of the pancreas with beta-cell antibodies in group III compared to group II. Conclusion Propolis showed protective and preservative effects on pancreatic beta-cell integrity in STZ-induced diabetic rats. This study suggests the need for further research to elucidate the exact mechanism of this effect.

HLA-haplotypes, anti- beta-cell antibodies: their role in prognostication of type 1 diabetes mellitus (results of 11-year observation)

The risk of type 1 diabetes mellitus (DM1) depends in the first place on the presence of predetermining HLA DR haplotypes, DQ genes, and specific autoantibodies.Aim. Prospective observation of DM1 risk groups (healthy sibs of DM patients) for the elucidation of the role of genetic and autoimmune abnormalities in thedevelopment of DM1. Materials and methods. Predetermining and protective haplotypes (HLA-DRBI, DQ genes) in combination with immunological markers (autoantibodiesto cytoplasmic structures of betbeta-cells (ICA), anti-insulin autoantibodies (IAA), anti-glutamate decarboxylase autoantibodies (GAD)) were studied in healthysibs of mean age 11.9?5.8 years selected from 143 discordant families of patients with DM1. In addition, a random sample of 599 DM1 patients (mean age7.5?6.2 years) was enrolled for molecular genetic studies (HLA-DRBI-DG genes). The control group comprised 200 subjects. Results. DQAI*0501-DQB1+0201 (DQ2) and DQAI*0301-DQBI*0302 (DG8) haplotype heterozygosity creating the highest risk of DM1, the healthy sibswere divided into 3 risk groups. The high risk group 1 (DQ2/DQ8) included 23 (13.5%) subjects, moderate risk group 2 (DQ2/X, DQ8/X) 85 (59.7%), andlow risk group 3 (X/X; X - any haplotype besides DQ2, DQ8) 63 (36.8%). The frequency of autoantibodies (AB) of 2 or 3 species in group 1 was higher thanin groups 2 and 3 (26.1, 14.1, 11.1% respectively, p&gt;0.05; 8.7, 2.4, 0%, p

Stiff-Man-Syndrome: Association of an autoimmune-encephalomyelitis with endocrinopathies and organ-specific antibodies

Stiff-Man-Syndrome (SMS) is a complex neuroimmunological disease of the Central Nervous System (CNS) with endocrinological, motoric, vegetative symptoms and is associated with autoantibodies to glutamate decarboxylase (GAD). SMS and its variants, stiff limb syndrome (SLS) and progressive encephalomyelitis with rigidity and myoclonus (PERM), appear to occur more frequently than hitherto thought. The cause of SMS is unknown. In a retrospective assessment of 92 patients (1989–2005) we investigated the correlation between SMS/SLS/PERM, associated autoimmune disorders, organ-specific antibodies and immungenetic HLA-association to find out the immunological differences between SMS, its variants and GAD-Ab positive- and negative patients. In total 54 of 92 patients with SMS and its variants were associated with 96 autoimmune diseases included autoimmune-endocrinopathies, e.g. Diabetes mellitus Typ I n =40, Hashimoto Thyroiditis n =13, Vitamine B12 Hypovitaminosis n =11, Pernicious anemia n =4 and Atrophic gastritis n =4. Associated immunopathies are most presented in patients with GAD-Ab: in 65 patients with GAD-Ab appeared 83 immunopathies, 26 patients without GAD-Ab only 13. In total 70% of autoimmune-endocrinopathies were diagnosed after the manifestation of this neurological disorder. Organ-specific antibodies are more frequent in patients with GAD-Ab than in patients without GAD-Ab, e.g. beta cell antibodies were found in 23 of 40 patients with GAD-Ab, but no antibodies were found in 13 patients without GAD-Ab. Two different groups can be distinguished pathogenetic. GAD-Ab as a Master-Ab is leading the way in the diagnostics of SMS.We have observed a noticeable association of this autoimmune-encephalomyelitis between GAD-Ab and HLA-haplotypes DQB1*03, DRB1*04, DQB1*02 and DQA1*0102. This show a similarity with diabetes mellitus Type I and may represent a genetic disposition.These results show interactions between neuroimmunology, endocrinology, immungenetics. Diagnostic, therapy and treatment of these patients indicate integrative and complementary proceedings. We hope that, e.g. natural medicine, ayurveda, TCM will help to find the cause of this disease and extend the therapeutical options in the treatment of Stiff-Man-Syndrome and many other neuroimmunological diseases.

Childhood obesity complicating the differential diagnosis of maturity-onset diabetes of the young and type 2 diabetes

To describe a proband with features of type 2 diabetes who was found to have concomitant maturity-onset diabetes of the young (MODY) and the consequent multigeneration genetic analysis. Familial genetic analysis. Tertiary university medical center. The proband was a 13.5-yr-old boy with marked non-ketotic hyperglycemia, obesity, systolic hypertension, and insulin resistance. His mother, maternal aunt, grandmother, and great grandmother had diabetes; his father was obese and had early ischemic heart disease. Clinical examination, laboratory work-up, and DNA study. Mutation in hepatocyte nuclear factor-1alpha gene, the most common cause of MODY. The proband showed elevated C-peptide level and was negative for beta-cell antibodies. On genetic analysis for MODY, the 291fsinsC mutation was identified in all affected family members. A younger sister who was obese but had no signs of impaired glucose tolerance was also tested on the basis of these findings and was found to have the same mutation. The patient, who presented with apparent type 2 diabetes, had concomitant MODY 3, inherited from his mother's side, and some features of type 2 diabetes secondary to marked obesity. This combination probably caused an earlier and more severe presentation of the disease and had significant implications for medical management. A search for MODY mutations should be considered in patients with a history of diabetes in three generations of one side of the family, even those in whom the clinical picture resembles type 2 diabetes.

Beta cell and insulin antibodies in treated and untreated diabetic cats

Beta cell and insulin antibodies are involved in the pathogenesis of diabetes in human patients. Beta cell antibodies have also been found in about 50% of newly diagnosed diabetic dogs. This study’s objective was to examine these antibodies’ role in feline diabetes. The serum of 26 newly diagnosed untreated diabetic cats, 29 cats on insulin therapy, 30 cats with diseases other than diabetes, and 30 healthy cats was examined for beta cell and insulin antibodies. For beta cell antibody testing, purified beta cells from a radiation-induced transplantable rat insulinoma were used. Serum from cats in which anti-beta cell antibodies were induced by injecting a purified beta cell suspension subcutaneously was used as a positive control. Following incubation with test sera, fluorescein-labeled anti-cat immunoglobulins were used to visualize binding between the beta cells and cat gamma globulins. Each serum was tested on two different tumor preparations. For the detection of insulin antibodies, a charcoal separation method was used. It was found that none of the healthy cats, none of the newly diagnosed, untreated diabetic cats and none of the cats with diseases other than diabetes had antibodies against beta cells or against endogenous insulin. Four diabetic cats (14%) that had been treated with different insulin preparations had insulin antibodies. It is concluded that immune-mediated processes are not causing diabetes in the cat. Further studies are needed to evaluate if antibodies directed against exogenous insulin alter the response of diabetic cats to insulin.

Gastric parietal cell antibodies are associated with glutamic acid decarboxylase-65 antibodies and the HLA DQA1*0501-DQB1*0301 haplotype in Type 1 diabetes mellitus. Belgian Diabetes Registry.

To assess the prevalence of thyrogastric autoimmunity in relation to age, sex, beta-cell antibody status and HLA DQ haplotypes in Type 1 diabetes mellitus. One hundred and seventy-one patients with Type 1 diabetes mellitus were studied (male/female 86/85; mean age 19 +/- 11 years; duration of diabetes 5 +/- 4 years). Islet cell antibodies (ICA) and parietal cell antibodies (PCA) were measured using indirect immunofluorescence; glutamic acid decarboxylase-65 antibodies (GADA) by radiobinding assay and thyroid peroxidase antibodies (TPO) with an immunoradiometric assay (IRMA). The majority of subjects (81.3%) showed one or more autoantibodies. The prevalence rates were: GADA 64.9%, ICA 46.2%, PCA 19.9% and TPO 19.3%. Patients with ICA+ > or = 3 years after diagnosis had a higher prevalence of GADA (P = 0.03, odds ratio (OR) 2.66) and thyrogastric antibodies (P = 0.05, OR 2.23) than subjects ICA- after 3 years. PCA+ patients were older (P = 0.04), had a higher prevalence of GADA (P = 0.005, OR 3.89) and TPO (P = 0.05, OR 2.50) than PCA- subjects. Logistic regression analysis showed that PCA status was determined by the HLA DQA1*0501-DQB1*0301 haplotype (beta = 2.94, P = 0.04) and GADA status (beta = 2.44, P = 0.041). Thyrogastric antibodies are highly prevalent in Type 1 diabetes mellitus, especially in patients with persisting ICA. Screening for gastric autoimmunity is particularly advised in patients who are positive for GADA and for the HLA DQA1*0501-DQB1*0301 haplotype.

Serological markers of autoimmunity in renal transplant patients before and after alpha-interferon therapy for chronic hepatitis C.

Chronic hepatitis C is the major cause of chronic liver disease after successful cadaveric renal transplantation. The aims of this prospective, open study were to assess in such a population, firstly the prevalence of different organ-specific and nonspecific antibodies and related disorders, and secondly their outcome after inteferon-alpha therapy as well as the incidence of new immunologic disorders under and after this therapy. In 15 cadaveric renal transplant patients (10 men, 5 women, ages 29-65 years) with chronic hepatitis C and histological features of chronic active hepatitis, undergoing chronic immunosuppression (ciclosporine A with or without steroid and azathioprine) and treated with recombinant alpha 2b-interferon (IFN alpha) (mean duration 142 +/- 35 days), we assessed before and after this therapy the serum levels of cryoglobulinemia, rheumatoid factors (RF), thyroid-stimulating hormone (TSH), free thyroxine (fT4), and antinuclear (ANA), antismooth muscle (ASMA), antimitochondrial (AMA), anti-LKM1, antimicrosomal thyroid (MCA), antithyroglobulin (TGA) autoantibodies. At the start of IFN alpha therapy, 14 of 15 patients had detectable autoantibodies (RF: 9; ANA > 1/50: 8; ASMA > 1/50: 4; other autoantibodies: 0); 1 had cryoglobulinemia. At the end of therapy the cryoglobulinemia had disappeared, the preexisting autoantibodies remained present in all patients but 2; 3 patients had developed MCA without evidence of clinical or biological thyroid abnormalities and 3 others had developed either RF (1) or ANA (1) or ASMA (1), without any related symptoms. One patient developed transient type II diabetes mellitus without anti-Langerhans beta-cell antibodies. Finally, the occurrence of autoantibodies in our patients was associated either with HLA DR3 or DR4 or DR7 phenotypes. We found that the prevalence of extrahepatic immunologic abnormalities was high in renal transplant patients with chronic hepatitis C and no exacerbation was observed during of after IFN alpha therapy. The most frequent autoantibody appearing after IFN alpha therapy was MCA although without thyroid abnormalities.

A qualitative assay for beta cell antibodies. Preliminary results in dogs with diabetes mellitus

Purified beta cells from a radiation-induced transplantable rat insulinoma were used to detect beta cell antibodies in serum from untreated diabetic dogs. Serum from dogs in which anti-beta cell antibodies were induced by injecting a purified beta cell suspension subcutaneously was used as positive control. Following incubation with test sera, fluorescein-labeled anti-dog immunoglobulins were used to visualize binding between the beta cells and dog gamma globulins. Nine of the 23 diabetic dogs showed a strongly positive reaction which was characterized by a ring fluorescence, three showed a weak reaction and 11 were negative, i.e. they showed diffuse fluorescence. In contrast, 14 of the 15 healthy dogs showed diffuse fluorescence and one dog showed a weakly positive reaction. Thyroid, liver and kidney cells did not elicit ring fluorescence. Although females (spayed and intact) represented the majority of the diabetic dogs, there was no correlation between sex and the occurrence of antibodies in the diabetic dogs. There was also no correlation to the age of the dogs. In conclusion, we have developed a specific test for anti-beta cell antibodies. The test is reproducible and economical to perform on a large number of samples.