Immunoassays have been used in recent years to detect the presence of infectious diseases. In order for the assay to be useful, it must detect a particular organism with a high degree of reliability. In most cases, this requires the isolation and reaction of antigens peculiar to the organism with corresponding antibodies. For the test to be commercially successful, it also needs to be relatively inexpensive, simple to use and rapid.
One such organism which can be detected by immunoassay is Chlamydia trachomatis (herein C. trachomatis) which is one of two microbial species of the genus Chlamydiaceae, order Chlamydiales. There are 15 or more strains of this species which are the causes of a number of human ocular and genital diseases including trachoma, inclusion conjunctivitis, lymphogranuloma venereum, nongonococcal urethritis and proctitis. Infection from C. trachomatis is pervasive in the general population so that it is believed that there are millions of cases each year of nongonococcal urethritis alone.
Despite the increasing control of various viruses by vaccination and various anti-viral agents, infection by organisms such as herpes simplex virus (HSV) remains a serious problem. There are two types of HSV, type 1 which occurs mainly around the mouth while type 2 occurs primarily around the genital area of the human body. Skin infections and viral encephalitis are but two of the serious results from HSV infection.
Gonorrhea is a disease usually transmitted by sexual contact caused by a bacterium of the Neisseria genus, especially N. gonorrhoeae. The disease has plagued mankind for thousands of years, and although antibiotics have helped control its spread, it still persists in epidemic proportions in many parts of the world. The importance of detection and treatment of this organism is well recognized. N. meningitidis and N. lactamica are also species of considerable medical and diagnostic interest.
Because of the widespread nature of these diseases, there is considerable interest in having a rapid, simple and reliable test for detection of chlamydial, gonococcal and herpes organisms. Considerable research has been carried out to find useful ways to extract detectable antigen from the organism. See for example, U.S. Pat. Nos. 4,427,782 (issued Jan. 24, 1984 to Caldwell et al) and 4,663,291 (issued May 5, 1987 to Rose) and E.P. Publications 174,106 (Becton) and 193,431 (Caldwell et al) which describe methods for extracting chlamydial antigens.
Extraction of antigen from the organisms in a biological specimen is critical to providing an accurate, rapid and sensitive assay. Many varied techniques have been used for extraction including physical disruption of the cells by sonication, heating or centrifugation. Chemical extraction compositions have also been developed. For example, the lipopolysaccharide antigen of C. trachomatis has been extracted using a mixture of phenol and water in E.P. Publication 193,431 (noted above).
Ethanolamine has been used in combination with surfactants and high temperature heating (for example, 70.degree.-110.degree. C.) to extract Chlamydial antigens as described in E.P. Publication 183,383 (IQ BIO). Extraction using high pH and heating is described in E.P. Publication 174,106 (noted above).
Assays for C. trachomatis and N. gonorrhoeae carried out using a solid support are described in U.S. Pat. Nos. 4,497,899 and 4,497,900 (both issued Feb. 5, 1985 to Armstrong et al and Abram et al, respectively). The described assays are performed by extracting antigen from the organism using a combination of an anionic or nonionic surfactant, deoxycholate and dithiothreitol, and coating it on a bare solid support. The coated antigen is then detected with either one or two antibodies, one of which is suitably labeled. Attachment of antigen is apparently achieved by incubating the coated support for an extended time sufficient to cause adsorption of antigen thereon (Col. 2, lines 51-55). From the examples of the patent, this time is determined to be at least 30 minutes at elevated temperature (37.degree. C.). For example, the assay described in U.S. Pat. No. 4,497,899 takes at least 3 hours to perform.
A significant advance in the art is described in copending U.S. Ser. No. 255,928, filed Oct. 7, 1988 by Pronovost, Mauck, Sullivan, Greer and Gilbert, and entitled "High pH Extraction Composition and Its Use to Determine a Chlamydial, Gonococcal or Herpes Antigen". The invention claimed and described therein provides a rapid and effective means for extracting chlamydial, gonococcal or herpes antigens without resort to extended heating. Rather, a pH above about 8 and an ethanolamine are used for effective antigen extraction.
Despite the important advance provided by the invention just described, whole blood or mucous is sometimes present in biological specimens and acts to interfere with antigen extraction. Moreover, the presence of blood or mucous often reduces the sensitivity of the assay even if the extraction of antigen is rapid and efficient. The extraction techniques described above do not effectively overcome the problems caused by the presence of blood or mucous in the specimen.
It would be desirable to have a much more rapid test for chlamydial, gonococcal or herpes organisms which has high reliability and can be performed at room temperature. It would also be desirable to efficiently extract antigen and provide an efficient assay thereof even if whole blood or mucous is present in the specimen.