title: Quantification of the major brown shrimp allergen Pen a 1 (tropomyosin) by a monoclonal antibody-based sandwich ELISA.
abstract: BACKGROUND
  Among 13 allergens found in extracts of cooked brown shrimp (Penaeus aztecus) the 36 kd muscle protein tropomyosin has been identified as the only major shrimp allergen (Pen a 1). Cross-reacting molecules with similar molecular weights were detected in other crustacea species such as crab, lobster, and crawfish. Because Pen a 1 and Pen a 1-like allergens are important in crustacea allergy, the aim of this study was to develop a monoclonal antibody (mAb)-based sandwich ELISA to quantify Pen a 1 and to evaluate Pen a 1 levels in four commercial shrimp, crab, and lobster extracts.
  
  
  METHODS
  Two Pen a 1-specific mAbs with different epitope specificities were selected. ELISA plates coated with captured mAb 3.2 were incubated with samples containing Pen a 1. Bound Pen a 1 was detected by a combination of biotinylated mAb 4.9.5 and alkaline phosphatase-labeled streptavidin.
  
  
  RESULTS
  The optimized sandwich ELISA could detect Pen a 1 concentrations ranging from 4 to 125 ng/ml. Four commercial shrimp extracts demonstrated a 40-fold difference in Pen a 1 levels (24 to 920 microg/ml). Crab and lobster extracts contained detectable levels of Pen a 1-like proteins. No reactivity to cockroach, house dust mite, oyster, codfish, or peanut extracts was detected, which indicates that the developed assay is crustacea-specific.
  
  
  CONCLUSIONS
  A sensitive sandwich assay was developed to quantify Pen a 1. This assay will be helpful to standardize shrimp extracts in regard to the content of the major allergen, Pen a 1, and to study cross-reactivities among and evaluate occupational exposure to different crustacea species.
authors.affiliation: Tulane University Medical Center, Department of Medicine. New Orleans, La. 70112, USA.
authors.forename: B J
authors.initials: BJ
authors.lastname: Jeoung
authors.forename: G
authors.initials: G
authors.lastname: Reese
authors.forename: P
authors.initials: P
authors.lastname: Hauck
authors.forename: J B
authors.initials: JB
authors.lastname: Oliver
authors.forename: C B
authors.initials: CB
authors.lastname: Daul
authors.forename: S B
authors.initials: SB
authors.lastname: Lehrer
mesh_terms: D000485:Allergens / Q000032:analysis* / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal; D003386:Brachyura / Q000737:chemistry / Q000276:immunology; D003429:Cross Reactions; D033364:Decapoda / Q000737:chemistry / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D008121:Nephropidae / Q000737:chemistry / Q000276:immunology; D012015:Reference Standards; D012044:Regression Analysis; D012680:Sensitivity and Specificity; D014335:Tropomyosin / Q000032:analysis* / Q000276:immunology
chemical_list: D000485:Allergens; D000911:Antibodies, Monoclonal; D014335:Tropomyosin