Source Of Antigenic Material Research Articles

Extracellular vesicles induce protective immunity against Trichuris muris.

Gastrointestinal nematodes, such as Trichuris trichiura (human whipworm), are a major source of morbidity in humans and their livestock. There is a paucity of commercially available vaccines against these parasites, and vaccine development for T. trichiura has been impeded by a lack of known host protective antigens. Experimental vaccinations with T. muris (murine whipworm) soluble Excretory/Secretory (ES) material have demonstrated that it is possible to induce protective immunity in mice; however, the potential for extracellular vesicles (EVs) as a source of antigenic material has remained relatively unexplored. Here, we demonstrate that EVs isolated from T. muris ES can induce protective immunity in mice when administered as a vaccine without adjuvant and show that the protective properties of these EVs are dependent on intact vesicles. We also identified several proteins within EV preparations that are targeted by the host antibodies following vaccination and subsequent infection with T. muris. Many of these proteins, including VWD and vitellogenin N and DUF1943‐domain‐containing protein, vacuolar protein sorting‐associated protein 52 and TSP‐1 domain‐containing protein, were detected in both soluble ES and EV samples and have homologues in other parasites of medical and veterinary importance, and as such are possible protective antigens.

Design and construction of a new recombinant fusion protein (2b2t+EPC1) and its assessment for serodiagnosis of cystic echinococcosis.

The immunodiagnostic tests for cystic echinococcosis (CE) are mostly serological tests based on ELISA that use hydatid cyst antigens for primary screening because of its simple preparation and availability. The challenge to develop new serological methods (as compared to those based on the hydatid cyst fluid antigens) to meet the gold standard remains. Appropriate sources of antigenic material are necessary for application to improve the efficacy of immunodiagnostic tests at a population level. In the current study, a fusion protein containing the coding sequence of antigen B2t and two sequences of EPC1 antigen with some modifications was reconstructed. Using bioinformatics tools, these sequences were joined together by applying the sequence of a rigid α-helix-forming linker to obtain an appropriate structure of a fusion protein. Synthetic recombinant fusion protein was expressed using pET28a as a vector and evaluated by indirect ELISA test for sera from patients with hepatic CE and other parasitic infections. The sensitivity of the fusion protein was lower (88.46%) than the available ELISA kit (96.15%). However, the differences in sensitivity were not statistically significant as compared to the recombinant fusion peptide with the commercial kit (p=0.269). The specificity of the recombinant fusion protein (95.45%) was not significantly lower than the commercial kit (96.59%; p=1.000). Moreover, surprisingly there was no difference in the cross-reactivity values of performance between the recombinant-ELISA and commercial kit. The positive and negative predictive values of the recombinant antigen were achieved as 92% and 93.33%, respectively, while for the commercial kit, they were obtained as 94.33% and 97.70%, respectively. In conclusion, as an early evaluation of these antigens the performance of our recombinant fusion protein in ELISA is relatively promising. Although, it seemed that this peptide with specific antigenic epitopes might be more appropriate for the serological evaluation of CE by use of bioinformatics tools, our findings showed that cross-reactions and a negative reaction could occur in clinical performance. This fusion protein may have utility for diagnosis in humans, but further evaluation is needed using the WHO ultrasound classification for CE.

A Study of Cross-Reactivity Between Recombinant EPC1 Antigen of Echinococcus granulosus in Serum from Patients with Confirmed Cystic Echinococcosis Infection and Other Parasitic Infections.

A standardized test for the serodiagnosis of cystic echinococcosis (CE) remains an important challenge because of the problems in specificity and sensitivity of the available commercial kits and lack of proper evaluation of antigen. Using appropriate sources of antigenic material is crucial in improvement of the serological methods such as enzyme-linked immunosorbent assay (ELISA). This study was conducted to evaluate the performance of protein named Echinococcus protoscolex calcium binding protein EPC1 for the detection of antibodies in sera from patients with CE. Expressed and purified recombinant protein EPC1 (rEPC1) was used as antigen in ELISA method. Characterization of the rEPC1 antigen was evaluated using the serum of 25 patients with both surgical and imaging confirmed CE and 25 healthy donors as negative controls. Also, a panel of sera including chronic toxoplasmosis (IgG positive), strongyloidosis, fascioliasis, toxocariasis, and kala azar were used and patients with related parasites were confirmed by medical laboratories or clinically by research centers using microscopy or specific ELISA. rEPC1 showed relatively promising performance in total IgG ELISA for the detection of antibodies in sera from the negative controls, and the cut off value 0.4 units of optical density at 490 nm was calculated for ELISA. In this study, sensitivity of 100%, specificity of 93.7, positive predictive value of 92.6%, and negative predictive value of 100% were calculated for rEPC1. On the other hand, commercial ELISA kit based on the native antigen B of Echinococcus granulosus had sensitivity of 96.2% and specificity of 96.8%. No significant difference was found for sensitivity or specificity between the rEPC1 and commercial kit. However, rEPC1 may be a valuable antigen for diagnosis of human CE.

Application of recombinant Echinococcus granulosus antigen B to ELISA kits for diagnosing hydatidosis

Echinococcus granulosus causes human cystic echinococcosis as an important public health problem in many regions of the world. There are some problems in primary diagnosis such as cross-reaction with sera from patients with other parasitic disease in serological tests. The use of an appropriate source of antigenic material is a very important and crucial point in the improvement of the serodiagnostic features such as enzyme-linked immunosorbent assay (ELISA) method. We expressed and purified recombinant AgB of Echinococcus granulosus and used as antigen in ELISA method. Serum samples were given from 36 cystic hydatid disease patients that have been confirmed by surgical operation as well as 36 healthy individuals sera were tested by ELISA method using recombinant AgB and compared with commercial kit (Euroimmun) for specificity and sensitivities value. The sensitivity of 91.66% and specificity of 97.22% were determined by homemade kit.

Immunologic analyses of bovine bone treated with a novel tissue sterilization process

This study evaluates the efficacy of removal of xeno-antigens from bovine bone using a patented BioCleanse process for decellularization of allograft tissues for clinical implantation. BioCleanse deploys a combination of chemicals and several high pressure rinses to achieve standardized sterility assurance levels. This method produces sterile grafts without reducing allograft bone biomechanical properties and effectively removes cells, lipids, and other sources of antigenic material from human allografts for clinical use. In this investigation, BioCleanse is evaluated for its potential in removing xenograft antigens from bovine bone grafts followed by immunologic evaluation in the subcutaneous pouch of immunocompetent rats. The alpha-galactosyl (alpha-gal) epitope with the structure Galalpha1-3Galbeta1-4GlcNAc-R constitutes a critical component of xenoantigens and its removal using BioCleanse from bovine bone was compared with tissue levels of unprocessed bone. The relative degree of antigen removal was also determined through measuring the pro-inflammatory cytokine, tumor necrosis factor (TNF)-alpha, and through the use of histologic grading of cellular infiltrates into bone. Compact cortical bone inhibited immune cell migration but cancellous bone demonstrated cellular increase and bone resorption in the untreated control group. The alpha-gal xenoantigen level was significantly lower in both cortical (P < 0.001) and cancellous bone (P < 0.001) compared with controls. TNF-alpha levels were significantly (P < 0.001) reduced compared with untreated controls when human acute monocytic leukemia cells were exposed to cortical or cancellous bone. BioCleanse effectively removed xenoantigens and inflammatory markers justifying a follow up study in primates to determine these benefits in a model that is primed with preformed xeno-antibodies responsible for hyperacute rejection in hard tissues.

Antigens for the immunodiagnosis of Echinococcus granulosus infection: An update

The taeniid tapeworm Echinococcus granulosus is the causative agent of the echinococcal disease, an important zoonosis with worldwide distribution. Accurate immunodiagnosis of the infection requires highly specific and sensitive antigens to be used in immunodiagnostic assays. The choice of an appropriate source of antigenic material is a crucial point in the improvement of the diagnostic features of tests, and must be based on the developmental stage of the parasite and the host. The most common antigenic sources used for the immunodiagnosis of echinococcal disease are hydatid cyst fluid, somatic extracts and excretory-secretory products from protoscoleces or adults of E. granulosus. Hydatid cyst fluid is the antigenic source of reference for immunodiagnosis of human hydatidosis, which is mainly based on the detection of antigens B and 5. Somatic extracts have been widely used in the serodiagnosis for E. granulosus infection in dogs and ruminant intermediate hosts, although in the last few years the detection of excretory-secretory products of the worm in faeces (coproantigens) have become the most reliable method for the detection of the parasite in the definitive host. This review emphasizes recent advances in the identification and characterization of novel antigens with potential for the immunodiagnosis of echinococcal disease. Progress in recombinant technologies and synthetic peptides are also discussed. The paper highlights the need to search for new antigenic components with high diagnostic sensitivity and specificity, a fact that remains a crucial task in the improvement of the immunodiagnosis of the disease.

Progressive polyarthritis induced in BALB/c mice by aggrecan from normal and osteoarthritic human cartilage

To find an "unlimited" source of antigenic material (aggrecan) for arthritis induction in BALB/c mice; to analyze the specificities of immune reactions to aggrecan and type II collagen in 2 arthritis-susceptible murine strains, BALB/c mice for proteoglycan (aggrecan)-induced arthritis and DBA/1j mice for collagen-induced arthritis; to compare the histopathologic features of arthritis induced by purified aggrecans or total extracts of osteoarthritic (OA) cartilage; and to determine arthritis susceptibility in various BALB/c colonies. Aggrecans from total extracts of human fetal, normal adult, OA, and rheumatoid cartilage samples and from osteophytes were isolated, purified by gradient centrifugation, deglycosylated, characterized, and tested for arthritis induction. Purified type II collagen and salt-soluble collagens from OA cartilage were denatured, stromelysin treated, and used for immunization and arthritis induction in arthritis-susceptible (DBA/1j and BALB/c) murine strains. Chondrocytes from OA cartilage synthesize predominantly fetal-type aggrecan, which is the most efficient antigenic material for arthritis induction in BALB/c mice. The critical autoimmune/arthritogenic T cell epitopes of aggrecan are located in the G1 domain. Although most of the aggrecan molecules are heavily degraded and lost from OA cartilage, the G1 domain-containing fragments accumulate in OA cartilage. The amount of G1-containing fragments is approximately twice as much in OA than in normal adult articular cartilage, and the arthritogenic epitope(s) remains intact in G1-containing fragments retained in cartilage. Thus, total extracts of OA cartilage (without additional purification), if deglycosylated appropriately, can be used as arthritogenic material in BALB/c mice. Predominantly G1 domain-containing fragments of aggrecan accumulate in OA cartilage, and these are the fragments which induce arthritis in BALB/c mice. Arthritis induction is highly specific for aggrecan epitopes and dictated by the genetic background of the BALB/c strain.

Status asthmaticus. Causes and management.

The life-threatening emergency represented by status asthmaticus points up the need for both specific and nonspecific forms of therapy in all cases of allergic asthma. Many of the common causes of death in this situation are iatrogenic. Among them are improper use of drugs, failure to remove sources of antigenic material (e.g., feather pillows), delay in institution of vigorous treatment, and premature discontinuance of therapy.