A Micro‐Neutralization System for Detection of s‐CRN Antibody to Herpes Simplex Virus

Abstract

It was learned that the ordinary micro-neutralization system with herpes simplex virus (HSV) gave a composite result of the initial neutralization and the effect of antibody on subsequent growth of unneutralized virus. In the case of slow-reacting complement-requiring neutralizing (s-CRN) antibody, which was detected by incubating virus-serum mixtures at 4 C for 3 days before addition of complement, the titer obtained was lower than expected from the result of the plaque reduction test. This was thought ascribable to its low ability to prevent viral breakthrough caused by growth of unneutralized virus. This was overcome by adding an appropriate amount of hyperimmune antibody at 3 hr after addition of cells. The endpoint of s-CRN antibody so determined was but slightly lower than that obtained by the plaque reduction test. Early (I-week) rabbit sera, which were negative in the ordinary micro-neutralization test, titered I: 2,560 to I: S, 120 when tested by this method. When the 3-day sensitization in the cold was substituted by 5-hr incubation at 37 C, the titer obtained was 2 to 4-fold lower; in this case, however, the whole process could be finished within 3 days. Also, s-CRN antibody reactive with type 2 HSV in homologous and heterologous sera could be detected by the same method using type I hyperimmune serum as the additional antibody.Routine procedures for virus neutralization did not use complement (C) in former days. However, in the case of herpes simplex virus (HSV), we found earlier that addition of C to serum dilutions enhanced the neutralization endpoint, the effect being especially marked with early immune sera (10, II). The antibody so detected was named C-requiring neutralizing (CRN) antibody. Furthermore, when virus-serum mixtures were held in the cold for I to 3 days before the addition of C, unexpectedly high endpoints were demonstrated. We ascribed this to presence of slow-reacting CRN (s-CRN) antibody (8,9,13). In an analysis of these phenomena, we postulated that the avidity of neutralizing antibody might be heterogeneous among molecules of IgG as well as of IgM, and the classical neutralization test detected only part of the whole neutralizing activity (13). It was further revealed that IgM s-CRN antibody required more C than did IgG s-CRN antibody, and was slower in sensitization reaction than the latter (13).In order to apply the detection of s-CRN antibody to routine practice, we 1037

Detection of Bovine Rhinotracheitis Virus Antibody by Neutralizing Test and ELISA in Experimentally Infected Rabbits

“…Although virus isolation and clinical signs were not observed in this experiment, these results suggest that the antibody response was produced in rabbits infected with IBR virus in agreement with the results reported for bovine (2,3). Addition of anti-rabbit IgG goat globulin (AIG) or complement to the virus-serum mixture increased sensitivity to the detection of neutralizing antibody, as reported by SATO et al (10) or YOSHINO and ABE (14). It is interesting that the titer of the antibody with complement became very high 1 week after the inoculation in both of the rabbit groups (Fig.…”

Analysis of herpes simplex virus type 1 restriction fragment length polymorphism variants associated with herpes gladiatorum and Kaposi's varicelliform eruption in sumo wrestlers

“…Neutralizing and complement-requiring neutralizing (CRN) antibodies against HSV-1 in serum specimens were titrated soon after the specimens were taken, as previously described (Asano et al, 1994;Yoshino & Abe, 1981;Yoshino & Isono, 1978). Complement (0.25 units) was added to each well of 96-well micro-test plates for CRN antibody titrations (Yoshino & Abe, 1981). RFLP analyses were performed as previously described Ozawa et al, 2006) using the restriction endonuclease maps of strain F as references (Hayward et al, 1975;Locker & Frenkel, 1979;Roizman, 1979; Roizman & Tognon, 1983).…”

“…The virus suspensions used in this study were prepared from virus stocks that had not been passaged more than five times after isolation. Neutralizing and complement-requiring neutralizing (CRN) antibodies against HSV-1 in serum specimens were titrated soon after the specimens were taken, as previously described (Asano et al, 1994;Yoshino & Abe, 1981;Yoshino & Isono, 1978). Complement (0.25 units) was added to each well of 96-well micro-test plates for CRN antibody titrations (Yoshino & Abe, 1981).…”