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Introduction: different  isotypes  of  antibody  can  be  produced  by  immune  system  after  antigen  contact.  Detection  and  measurement  of  different  classes  of  antibody  against  the  antigen  is  very  important  in  some  cases.  The  aim  of  this  study  is  designing  of  an  ELISA  method  on  the  basis  of  inhibition  of  enzyme  activity  by  using  a  non-competitive  inhibitor.  Therefore  in  this  study  rheumatoid  factor  is  used  as  a  model  for  the  detection  of  different  other  classes  of  antibodies  against  the  antigen.
Materials  and  Methods:  In  this  cross  sectional  analytical  study, we  measured  IgM  and IgA   rheumatoid  factors  in  sera  of  10  patients  with  rheumatoid  arthritis  and  positive  latex  test , by  mixed  and  routine  ELISA.  In  mixed  ELISA  the activity  of  the  first  conjugated  enzyme  was  blocked  by  a  non-competitive  inhibitor  after  adding  the  substrate. Then  the  next  conjugated  antibody, which  was  specific  for  another  isotype, was  added. By  optical  density, results  was  comparisoned  with  routine  ELISA.
Results:  The  obtained  results  showed  that  the  average  optical  density  is  lower  when  compared  with  routine  ELISA , but  the  difference  is   not  statistically  significant.  however  these  two  methods  did  not  show  any  significant  difference  in  quantifying  antibody  isotypes. Also  there  is  a  positive  association  between  mixed  and  routine  ELISA (r=0.9, p=0.001).
Discussion: Lower  optical  density  in  mixed  ELISA  is  probably  because  of  stick  hindrance  by  the  first  conjugate. So, because  there  is  no significant  difference  between  the  results  of  these two  types  of  ELISA, and  also  no  need  to  repeat  the  test  for  each  isotype  in  this  method, it  is  recommended  to  use the  new  method  instead  of  the  routine  one  to  save  time  and  reagents.
 

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Background: Enterotoxigenic Escherichia coli bacterium is the most important bacterial agent causing diarrhea. Specific virulence factors, such as enterotoxins and colonization factors, distinguish ETEC from other classes of diarrheagenic E.coli. In this study, heat-labile toxin was purified which could be utilized for anti-toxin assay in GM1 gangelioside receptor-ELISA based method and for identification of ETEC producing toxin. Materials and Methods: In this experimental study, bacterial strain producing heat-labile toxin was first cultivated for production and purification of toxin. Then supernatant soluble proteins were precipitated with ammonium sulfate and purified using biochemical methods. Finally, purified protein was dialyzed against Tris 0.02 mM pH 8 and analyzed on gel electrophoresis. GM1 gangelioside receptor-ELISA based method was used for detection and assessment of the purified toxin. Through this method, the effect of anti-recombinant heat-labile toxin B subunit neutralization on heat-labile toxin was investigated. Results: Toxin purification was revealed by the presence of 12 and 28 KD protein bands. This study demonstrated that anti-recombinant heat-labile toxin B subunit antibody can detect the purified toxin and can inhibit its binding to GM1 receptor up to 80%. Conclusion: Purification of heat-labile toxin and gangelioside receptor-ELISA assay can be used for accurate detection and epidemiological study of clinical isolates.

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Background: The present study aimed to investigate the immunomodulatory activity of molecules secreted by decidual cells on dendritic cells (DCs) function in abortion-prone compared with non-abortion-prone mice.

Materials and Methods: The decidual cell supernatants (DS) were obtained from abortion and non-abortion mouse models. DCs were purified from CBA/J mice spleens and treated with antigen and DS. Treated DCs were injected into mice palms. After 5 days, draining lymph nodes were removed, cultured in the presence of cognate antigen, and proliferation of lymphocyte cells was measured by 3H-thymidin incorporation.

Results: Our results showed that immunosuppressive activity of DS from non-abortion-prone mice significantly decrease dendritic cells' ability to stimulate lymphocytes proliferation compared with DS from abortion-prone mice (Simulation index (SI) of 4.93 ± 0.34 versus 11.84 ± 0.79).We, also found that DS prepared from non-resorption sites compared with DS from resorption sites in abortion-prone mice had increased immunosuppressive activity on DC function (SI of 7.31 ± 1.02 versus 2.67 ± 0.49).

Conclusion: Due to our results, we concluded that immunomodulatory activity of molecules secreted within decidual tissue is different between abortion-prone and non-abortion-prone mice. Based on the key role of DCs in inducing fetomaternal tolerance, we claimed that these molecules, through modulation of DCs function, play crucial role on pregnancy outcome.