Efficacy of a recombinant Norwalk virus protein enzyme immunoassay for the diagnosis of infections with Norwalk virus and other human “candidate” caliciviruses

Abstract

A recombinant Norwalk virus antigen enzyme immunoassay (rNV EIA) for the detection fo Norwalk virus (NV) antibodies was evaluated using sera from volunteers and patients infected with Norwalk, Hawaii, Snow Mountain Agent, small round structured viruses (SRSV), and human calicivirus (HuCV) strains. The results for NV antibodies correlated well with previous results obtained using a NV radioimmunoassay. The test rNV EIA was found to be highly specific for NV. The results of tests on sera obtained from patients involved in confirmed outbreaks of SRSV in the UK showed that only 1/12 was due to Norwalk virus. Further evidence is presented that SRSV strain UK2 is antigenically related to NV.

An epidemiological investigation of Norwalk virus infection in South Africa

“…The EIA system of Graham and colleagues [14], as modified by Parker and colleagues [16], was used to detect IgG antibodies to NV. In the assay system horseradish peroxidase (HRP) conjugated rabbit antihuman IgG (Dako immunoglobulins, Denmark) was used with 3,3',5,5'-tetramethylbenzidine dihydrochloride (TMB) (Kirkegaard & Perry, Gaithersburg, MD) as substrate.…”

“…Samples were tested at a dilution of 1/1000 and wells giving an OD value > 01 and at least double the negative control were considered to be positive. A titre of < 1000 in the rNV EIA has been found to be equivalent to < 50 in the radioimmunoassay (RIA) and was therefore considered to be negative [16].…”

“…Limited epidemiological studies, carried out with antigens and antisera from human volunteer studies, have shown that in lessdeveloped areas such as Bangladesh, NV is contracted early in life [10], whilst in the USA it was an uncommon childhood infection [11]. The recent cloning and sequencing of the entire NV genome [12], and the consequent production of recombinant NV antigen (rNV) [13], has facilitated the development of sensitive and specific enzyme immunoassays (EIA) for the detection of NV antigen and antibodies [14][15][16]. In addition, the application of the reverse transcriptase polymerase chain reaction and subsequent molecular characterization of these viruses has enabled many SRSVs, including NV, to be classified within the Caliciviridae [17].…”

Measurement of IgA responses following Norwalk virus infection and other human caliciviruses using a recombinant Norwalk virus protein EIA

“…The recent success in sequencing the entire NV genome [2,3] and subsequent expression of self assembling NV capsids in baculovirus [4] has enabled enzyme immunoassays (EIA) to be developed to measure antibody responses to NV [5]. Previous studies with human reagents indicated that anamnestic responses to NV occurred in some individuals infected with other strains of small round structured viruses (SRSV) [6] and caliciviruses (HuCV) [7].…”

“…Previous studies with human reagents indicated that anamnestic responses to NV occurred in some individuals infected with other strains of small round structured viruses (SRSV) [6] and caliciviruses (HuCV) [7]. The recombinant NV enzyme immunoassay (rNVEIA) has been shown to be highly sensitive and specific for NV [5] but slight IgG responses to NV were demonstrated in volunteers challenged with Hawaii and Snow Mountain agent when sera were tested at low dilutions [8]. In contrast IgA responses only occurred in the volunteers challenged with NV [8].…”