Details:

Alternative Title:
J Immunol Methods
Personal Author:
Mhike, Morgen ; Hettick, Justin M. ; Chipinda, Itai ; Law, Brandon F. ; Bledsoe, Toni A. ; ... More +
Mhike, Morgen ; Hettick, Justin M. ; Chipinda, Itai ; Law, Brandon F. ; Bledsoe, Toni A. ; Lemons, Angela R. ; Nayak, Ajay P. ; Green, Brett J. ; Beezhold, Donald H. ; Simoyi, Reuben H. ; Siegel, Paul D. Less -
Description:
Diisocyanates (dNCOs) are low molecular weight chemical sensitizers that react with autologous proteins to produce neoantigens. dNCO-haptenated proteins have been used as immunogens for generation of dNCO-specific antibodies and as antigens to screen for dNCO-specific antibodies in exposed individuals. Detection of dNCO-specific antibodies in exposed individuals for diagnosis of dNCO asthma has been hampered by poor sensitivities of the assay methods in that specific IgE can only be detected in approximately 25% of the dNCO asthmatics. Apart from characterization of the conjugates used for these immunoassays, the choice of the carrier protein and the dNCO used are important parameters that can influence the detection of dNCO-specific antibodies. Human serum albumin (HSA) is the most common carrier protein used for detection of dNCO specific-IgE and -IgG but the immunogenicity and/or antigenicity of other proteins that may be modified by dNCO in vivo is not well documented. In the current study, 2,4-toluene diisocyanate (TDI) and 1,6-hexamethylene diisocyanate (HDI) were reacted with HSA and human hemoglobin (Hb) and the resultant adducts were characterized by (i) HPLC quantification of the diamine produced from acid hydrolysis of the adducts, (ii) 2,4,6-trinitrobenzene sulfonic acid (TNBS) assay to assess extent of cross-linking, (iii) electrophoretic migration in polyacrylamide gels to analyze intra- and inter-molecular cross-linking, and (iv) evaluation of antigenicity using a monoclonal antibody developed previously to TDI conjugated to Keyhole limpet hemocyanin (KLH). Concentration-dependent increases in the amount of dNCO bound to HDI and TDI, cross-linking, migration in gels, and antibody-binding were observed. TDI reactivity with both HSA and Hb was significantly higher than HDI. Hb-TDI antigenicity was approximately 30% that of HSA-TDI. In conclusion, this data suggests that both, the extent of haptenation as well as the degree of cross-linking differs between the two diisocyanate species studied, which may influence their relative immunogenicity and/or antigenicity.
Subjects:
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1,6-hexamethylene Diisocyanate 2,4-toluene Diisocyanate Article Cross-Linking Reagents Enzyme-Linked Immunosorbent Assay Haptenation Haptens Hemoglobins Humans Isocyanates Occupational Asthma Serum Albumin Toluene 2,4-Diisocyanate
Source:
J Immunol Methods. 431:38-44.
Pubmed ID:
26853746
Pubmed Central ID:
PMC4792703
Document Type:
Journal Article
Funding:
CC999999/Intramural CDC HHS/United States ; Y01 ES000001/ES/NIEHS NIH HHS/United States ; IAG#Y1-ES-0001-12/ES/NIEHS NIH HHS/United States
CC999999/Intramural CDC HHS/United States ; Y01 ES000001/ES/NIEHS NIH HHS/United States ; IAG#Y1-ES-0001-12/ES/NIEHS NIH HHS/United States Less -
Collection(s):
CDC Public Access
Main Document Checksum:
[+]
urn:sha256:94a5b49f740ae04245d7f229b7d853600d0877d904944f4491bd605977a61f30
Download URL:
https://stacks.cdc.gov/view/cdc/38493/cdc_38493_DS1.pdf
File Type:

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