The development and application of a blocking ELISA kit for the diagnosis of infectious coryza

Abstract

A blocking enzyme-linked immunosorbent assay (B-ELISA) kit for the diagnosis of infectious coryza was developed in this study. The kit was based on a recently described blocking ELISA that uses monoclonal antibodies to achieve specificity for antibodies to either Haemophilus paragallinaru m serovar A or serovar C. The results showed that the B-ELISA kit detected 96 and 90%, respectively, of chickens vaccinated or challenged with H. paragallinaru m serovar A. When used on chickens vaccinated or challenged with H. paragallinaru m serovar C, the kit detected 77 and 40%, respectively, as positive. The majority of sera from vaccinated chickens were still positive on the serovars A and C ELISAs 4 months after vaccination. Based on pen trial data, the serovar A B-ELISA kit had a sensitivity of 95% and a specificity of 100%. The serovar C B-ELISA kit had a sensitivity of 73% and a specificity of 100%. A range of field sera was examined with the kit, generating results that correlated with the known vaccination/disease history of the flocks examined. As freeze drying the monoclonal antibodies and the conjugate had some effect on optimal working concentration, the kit used liquid solutions of these two reagents. The kit could be stored for 7 days at 37°C, 10 months at 4°C and more than 1 year at -20°C. Our results suggest that the kit would be a useful aid in the diagnosis of infectious coryza in China and other countries where H. paragallinaru m serovars A and C are the predominant or sole serovars.

Development of a Blocking ELISA for Detection of Serum Neutralizing Antibodies against Newly Emerged Duck Tembusu Virus

“…Blocking ELISA, a specific method for antibody detection, has been used widely to diagnose human diseases [18], monitor animal infectious diseases [19], [20], [21], [22], [23], and detect viral antibodies in clinics or laboratories [24], [25], [26], [27], [28], [29]. The distinct advantages of blocking ELISA include high-volume sample testing, applicability across multiple species, and greater objectivity than some traditional techniques.…”

Seroprevalence of Mycoplasma gallisepticum in Commercial Broiler Chickens in Duhok Governorate

“…Serology is the best tool for detection of clinical and subclinical MG infection in poultry flock [24]. Enzymelinked immunosorbent assay is one of the most widely used serological tests that are broadly used for finding of Mycoplasma gallisepticum (MG) infections [17,25], since ELISA and HI tests only identify IgG antibodies that arise after 7-10 days (DPI) and last for 180 days [26,27].…”

Development of Elisa Test Kit for Antibody Against Avibacterium Paragallinarum

“…Moreover, there is no commercial test kit for HI method. The alternative method is the monoclonal antibodybased blocking enzyme-linked immunosorbent assay (blocking ELISA) (Miao et al, 2000;Sun et al, 2 0 0 7 ) but it is mainly used in research. Although blocking ELISA using monoclonal antibody shows high specificity and acceptable sensitivity (Zhang et al, 1999), there are some limitation in testing such as (1) monoclonal antibody have only for Page serovar A and C (2) monoclonal antibody are not commercial available ( 3) some isolates of A. paragallinarum do not react with monoclonal antibody (Blackall, 1999).…”

“…Uncertainty of testing results depends on individual skills and method of preparation. The alternative method is monoclonal antibody-based blocking enzyme-linked immunosorbent assay (blocking ELISA) (Miao et al, 2000;Sun et al, 2007) which is highly specific with acceptable sensitivity (Zhang et al, 1999). They reported that the sensitivity of blocking ELISA against infectious coryza for serovar A and serovar C were 78.7% and 64.7%, respectively, and the specificity of blocking ELISA against infectious coryza for serovar A and C were 99.…”