An immunogold assay system for the detection of antigen or antibody

A colloidal gold-based semi-quantitative manual immunoassay method for the detection of antibody or antigen has been developed. The colloidal gold particles are coated with an organic reagent which, in turn, is attached to the antibody by covalent bonds. The antibody or the antigen are immobilized on simple chromatography paper. The paper strips are developed with an appropriate immunogold reagent in a test tube in the presence of urine (Pregnancy or Ovulation test) or serum (Rubella test). The mixture migrates up the strips towards the test band. A purple band develops which indicates the presence of the corresponding antigen (Pregnancy, Ovulation test) or antibody (Rubella test). In this format, 50 mIU or more of hCG in urine can be detected in 5 minutes or less, and antibody to Rubella virus at an HAI titer equivalent of 8 or above in serum can be detected in 10 minutes or less. These test kits, known as Result Plus test systems, are simple, rapid, highly sensitive, and require no instruments to perform.     

Ophthalmic genetics practice and research in India: Vision in 2020

Ophthalmic genetics is a much needed and growing area in India. Ethnic diversity, with a high degree of consanguinity, has led to a high prevalence of genetic disorders in the country. As the second most populous country in the world, this naturally results in a significant number of affected people overall. Practice involves coherent association between ophthalmologists, genetic counselor and pediatricians. Eye genetics in India in recent times has witnessed advanced research using cutting edge diagnostics, next generation sequencing (NGS) approaches, stem cell therapies, gene therapy and genomic editing. This article will highlight the studies reporting genetic variations in the country, challenges in practice, and the latest advances in ophthalmic genetic research in India.     

Antigen-specific T cell receptor antagonism by antigen-presenting cells treated with the hemolysin of Listeria monocytogenes: a novel type of immune escape

We have examined the influence of listeriolysin O (LLO), the hemolysin secreted by the pathogenic bacterium Listeria monocytogenes, on major histocompatibility complex class II-dependent T cell activation. Stimulation of T cells by native antigens but not by peptides is inhibited upon pretreatment of antigen-presenting cells (APC) with LLO. Experiments presented here reveal that this inhibition is not due to a lack in processing of antigen by APC but is the result of an irreversible inactivation of T cells that recognize antigen on LLO-treated APC. Incubation of mixtures of two different T cells where only one antigen was presented on LLO-treated APC suggested that T cell inactivation is antigen specific. The inactivation was dominant and could be observed even in the presence of amounts of synthetic peptides that normally lead to T cell responses. This condition is reminiscent of the T cell inhibition observed when antagonistic and stimulatory peptides are added to APC at the same time. Our results thus reveal a novel type of interference by pathogens with antigen presentation and T cell stimulation that could give the pathogen a decisive advantage in dissemination and disease.     

Structural and functional anatomy of the globular domain of complement protein C1q

C1q is the first subcomponent of the classical pathway of the complement system and a major connecting link between innate and acquired immunity. As a versatile charge pattern recognition molecule, C1q is capable of engaging a broad range of ligands via its heterotrimeric globular domain (gC1q) which is composed of the C-terminal regions of its A (ghA), B (ghB) and C (ghC) chains. Recent studies using recombinant forms of ghA, ghB and ghC have suggested that the gC1q domain has a modular organization and each chain can have differential ligand specificity. The crystal structure of the gC1q, molecular modeling and protein engineering studies have combined to illustrate how modular organization, charge distribution and the spatial orientation of the heterotrimeric assembly offer versatility of ligand recognition to C1q. Although the biochemical and structural studies have provided novel insights into the structure-function relationships within the gC1q domain, they have also raised many unexpected issues for debate.     

HLA haplotypes and microsatellite polymorphisms in and around the major histocompatibility complex region in a Native American population with a high prevalence of scleroderma (systemic sclerosis)

Choctaw Native Americans in southeastern Oklahoma have the highest prevalence of scleroderma or systemic sclerosis yet found (469/100,000). An Amerindian HLA DR2 haplotype (DRB1*1602) was significantly associated with scleroderma in this population in a previous study. It is not known, however, if other disease genes are linked to this HLA haplotype. The regions flanking the HLA loci were studied with polymorphic microsatellite markers. An extended HLA DR2 (DRB1*1602, DQA1*0501, DQB1*0301, DPB1*1301) haplotype that includes the class I and III regions was identified which was significantly associated with scleroderma in the Oklahoma Choctaw. No other significant associations with microsatellite marker alleles immediately flanking the HLA region were found.     

Cross-protection against four species of chicken coccidia with a single recombinant antigen

A cDNA clone, SO7', from an Eimeria tenella cDNA library was inserted into the high-expression vector pJC264 and was expressed in Escherichia coli as a fusion protein, CheY-SO7', with a molecular mass of approximately 36 kDa. By using the purified recombinant antigen to immunize young chicks, it was demonstrated that a single dose, without adjuvant, not only protected against severe coccidiosis induced by infection with E. tenella but also protected chicks challenged with the heterologous species Eimeria acervulina, E. maxima, and E. necatrix. By using rabbit antiserum raised against recombinant CheY-SO7', Western blot (immunoblot) analysis of sporulated oocysts of all seven major species of chicken coccidia showed that all species tested contained proteins characteristic of the B class of antigens, of which CheY-SO7' is representative. It seems likely that a single B antigen could protect chickens against severe coccidiosis caused by infection with any of these Eimeria species. Although chicks exposed to prolonged, natural infection develop antibodies to B antigen, active immunization of young chicks with a protective dose of CheY-SO7' does not elicit a humoral antibody response, suggesting that the partial protection results from cell-mediated effector mechanisms. In addition, the cross-protective nature of the immunity indicates that the response to B antigen is different from that induced by natural infection, which elicits a species-specific immunity. To date, the protection induced by B antigen immunization, although remarkable for a single recombinant protein, is not sufficient to compete with prophylactic chemotherapy.     

Using daily ratings to confirm premenstrual syndrome/late luteal phase dysphoric disorder. Part I. Effects of demand characteristics and expectations.

Daily symptom ratings were evaluated in 31 women who met DSM III-R criteria for a diagnosis of Late Luteal Phase Dysphoric Disorder (LLPDD), and 34 women who reported no premenstrual symptomatology. Women in both groups were randomly assigned to be aware or unaware of the menstrual cycle focus of the study. Results revealed a consistent pattern of cyclic effects, with higher ratings in the mid-luteal, premenstrual and menstrual phases for many symptoms. Ratings of women reporting severe symptoms were not affected by knowing the purpose of the study and were higher than ratings in the asymptomatic group for many symptoms relevant to the LLPDD diagnosis. Awareness of the study focus did increase cyclicity in the ratings of asymptomatic women aware of the study focus compared with those who were unaware, but these differences were small and most were not significant. The implications of these findings for research and clinical diagnosis of PMS/LLPDD are discussed.     

Gastric pull-up by the retrosternal route for esophageal replacement: Feasibility in a limited-resource scenario

ObjectivesThe authors herein report the results of esophageal replacement by gastric pull-up technique through the retrosternal route as an option for esophageal replacement in a resource-constrained setup.MethodProspectively collected data upon twenty-two consecutive patients (male:female = 17:5) with mean age 24.9 months (7 months–12 years) and mean weight 7.9 kg (4.2–32 kg) who underwent retrosternal gastric pull-up for esophageal atresia (n = 18; 16 atresia with distal fistula & 2 pure atresia) and corrosive injuries to the esophagus (n = 4) over the past 8 years are presented.The management protocol and surgical technique have been described.Observations parameters included indication for esophageal replacement, age at surgery, sex of the child and other demographic details, clinical and operative findings, post-operative outcomes and follow-up details.ResultsRetrosternal gastric pull-up could be performed in all cases with no mortality or graft loss. Of 22, 20 cases were extubated on-table and 2 cases were extubated within 48 hours of surgery.Mean operative duration was 265 min (range: 175 min to 310 min) and blood loss was 115.3 ml (range: 80–400 ml). Dense vascular adhesions in the region of the esophageal hiatus were encountered in patients with abdominal esophagostomy (n = 4) which were probably related to the local dissection at the time of previous surgery.Minor anastomotic leak was observed in 8 of 22 patients which settled spontaneously over 21 days mean period (range: 18 to 31 days). Antegrade dilatation was required in 3 of 8 cases with minor leak. None of them required revision of anastomosis.Mean follow-up duration is 63 months (range: 11 months – 94 months). Weight gain after surgery was close to or beyond the 25th centile. Symptoms of dumping syndrome or GER were not observed in our cohort.ConclusionOur data have demonstrated the safety and feasibility of esophageal replacement by gastric transposition through the retrosternal route in a resource-limited setup. No significant difference has been observed from the results and complications reported in literature for the same procedure.Type of studyProspective observational study / treatment study.Level of evidenceLevel III.