Identification of low molecular weight allergens of American cockroach and production of monoclonal antibodies.
BACKGROUND
Previous studies have identified two allergic fractions (Cr-PI and Cr-PII) from crude American cockroach extract, from which the 72- and 78-kilodalton (kD) components of Cr-PI have been identified as major allergens.


OBJECTIVE
The objective of this study was to identify important allergens in Cr-PII and produce monoclonal antibodies.


METHODS
Sera of cockroach-sensitive atopic patients were examined for IgE binding to Cr-PII proteins and monoclonal antibodies were generated by fusion between spleen cells of BALB/c mice immunized with Cr-PII and FO cells.


RESULTS
Immunoblotting revealed that eight allergenic compounds in Cr-PII are able to bind specific IgE. Allergens of 18, 28, 32 and 45 kD bound 16.7% (2/12), 100% (12/12), 83.3% (11/12), and 100% (12/12) of the atopic sera tested, respectively. Cell fusion resulted in three stable subclones secreting monoclonal antibodies, and none of the monoclonal antibodies recognized any epitopes of German cockroach and Cr-PI of American cockroach. One monoclonal antibody reacted strongly with the 28- and 32-kD allergens of Cr-PII, and two monoclonal antibodies were able to bind the 18- and 12-kD components of Cr-PII with similar epitope specificities. All the epitopes defined by monoclonal antibodies are recognized by human IgE as demonstrated by a fluoroallergosorbent test (FAST) inhibition assay. Five commercial cockroach extracts were compared for Cr-PII levels by monoclonal antibody-base ELISA, and all extracts were found to contain detectable Cr-PII.


CONCLUSIONS
The 28-, 32-, and 45-kD components of Cr-PII may be considered as additional important allergens of American cockroach and monoclonal antibodies can be used to identify and define American cockroach allergens.
Wu
C H
CH
Department of Medical Research, Taichung Veterans General Hospital, Taiwan, Republic of China.
Hsieh
M J
MJ
Huang
J H
JH
Luo
S F
SF
D000485:Allergens / Q000032:analysis* / Q000737:chemistry / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry; D000918:Antibody Specificity; D001666:Binding Sites, Antibody; D000939:Epitopes / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000737:chemistry; D051379:Mice; D008970:Molecular Weight; D010526:Periplaneta / Q000276:immunology*; D012882:Skin Tests / Q000009:adverse effects
D000485:Allergens; D000911:Antibodies, Monoclonal; D000939:Epitopes; D007073:Immunoglobulin E