Immunoblotting (WB) & Transmission Electron Microscopy (TEM) - strategies used in diagnosis of human Infectious diseases

Abstract

Infectious diseases are a significant burden on public health and economic stability of societies all over the world. They have been among the leading causes of death and disability and presented growing challenges to health security and human progress for centuries. Infectious diseases are generally caused by microorganisms. The routes of them entry into host is mostly by the mouth, eyes, genital openings, nose, and the skin. Damage to tissues mainly results from the growth and metabolic processes of infectious agents intracellular or within body fluids, with the production and release of toxins or enzymes that interfere with the normal functions of organs and/or systems [1]. Advances in basic science research and development of molecular technology and diagnostics have enhanced understanding of disease etiology, pathogenesis, and molecular epidemiology, which provide basis for appropriate detection, prevention, and control measures as well as rational design of vaccine [2]. The diagnosis of infectious diseases is particularly critical for the prevention and control of the epidemic. Here we introduce the insights and detection methods of infectious disease, aiming to provide some helps for clinical diagnosis as well as epidemic prevention and control of infectious diseases.

Immunoblotting (WB) & Transmission Electron Microscopy (TEM) - strategies used in diagnosis of human Infectious diseases

Immunoblotting technique detects specific viral proteins isolated from a cell, tissue, organ, or body fluid. The development of sensitive and specific tests for human immunodeficiency virus type 1 (HIV-1) progressed rapidly after this retrovirus was found to be responsible for causing AIDS [39]. Immunoblotting has been one of the reference confirmatory tests for the diagnosis of HIV infection or after inconclusive enzyme immunoassay (EIA) results. Although difficulty in interpretation of immunoblotting results and the cost led to a reduction in overall use of WB technique, nevertheless immunoblots are still commonly used for various purposes, including clinical diagnosis of HIV-1, seroprevalence surveys, and for blood-donor screening. In addition, immunoblot assays have been used to confirm the anti-hepatitis C virus (HCV) reactivity [40]. In recent years immunoblotting has been established as an important prerequisite for the functional studies to understand protein composition of the purified viral particles, since it allows the analysis of specific proteins which result in better understanding of the infection process and the pathogenesis of viruses [41,42].

Most viruses are very small to be seen directly under a light microscope, and therefore could only be viewed with TEM (transmission electron microscopy). In 1948, smallpox and chicken pox were first differentiated by TEM [43] and thereafter early virus classifications depended heavily on TEM analysis. In particular many intestinal viruses were discovered by negative staining TEM microscopy [44, 45]. Although TEM has gradually been replaced by more sensitive methods such as PCR, nevertheless it still remains essential for several aspects of virology including discovery, description and titration of viruses. One of the major advantages of using TEM is that it does not require virus-specific reagents; this is of particular importance in an outbreak setting where the etiologic agent is unknown and therefore specific reagents may not be available to determine correct detection tests. Negative stained TEM technique continues to be a valuable tool for the discovery and identification of novel viruses including Ebola virus, henipavirus (Hendra and Nipah) and SARS [46-50]. A human monkeypox outbreak was detected in the US by TEM [51]. Nevertheless, due to the high instrument cost and the amount of space and facilities required, TEM is still only available in certain facilities.

Figure 5. Transmission Electron Microscopy of hantavirus virions^[52]

Summary

Infectious diseases are a real public health threat, outbreaks can have serious social, political, and economic effects. A complex number of factors relating to human behavior and activities, pathogen evolution, poverty, and changes in the environment as well as dynamic human interactions with animals have been found to contribute to infectious disease emergence and transmission. Aggressive research is warranted to unravel important characteristics of pathogens necessary for diagnostics, therapeutics, and vaccine development. Here we describe some strategies for the diagnosis of human infectious diseases, hoping to be helpful for clinical diagnosis and epidemic prevention and control of infectious diseases. To date, multiple diagnostic techniques have been developed. Various diagnostic tools show both significances and limitations. Conventional approaches to quantify infective viral particles are labor-intensive, time-consuming, and often associated with poor reproducibility. Immunological tests generally provide quick results, however, is quite expensive due to the requirement of antigen-specific antibody. While RT-PCR may be able to provide results within a matter of hours, it is laborious, requires a skilled operator, and is sensitive to contamination. TEM-based quantification, although highly accurate in determining the shape and the total number of viral particles, often considered time-consuming, extremely expensive and impractical for high sample numbers. Moreover, TEM sample preparation is tedious, and the technique requires sophisticated instrument and a skilled operator. To alleviate these limitations, there is still a need to develop new cost-effective analytical methods that can allow users to quickly and easily determine virus concentrations and reduce constrictions coupled with current assays. Nevertheless, any such emerging methods must be carefully evaluated in terms of their efficiency, precision and linear range. The evaluation of each diagnostic technique and approval from the FDA are necessary before practical application.

Products list: paired diagnostics grade antibodies (monoclonal antibody, mab) and antigens for infectious disease rapid test kit

Cytokine release syndrome (CRS) is an acute systemic inflammatory syndrome characterized by fever and multiple organ dysfunction.

Genemedi produces core diagnostic ingredients for test of infectious diseases and related syndrome.

GeneMedi offers paired diagnostics grade antibodies (monoclonal antibody,mab)and antigens for infectious disease rapid test kit including infection of HIV/AIDS,Hepatitis C virus(HCV), Influenza A/B,Treponema Pallidum caused Syphilis,Helicobacter pylori (H. pylori) Bacteria, Herpes simplex virus 1/2(HSV-1/HSV-2),Cytomegalovirus (CMV),rubella virus,toxoplasma gondii and so on.

All the antibodies of inflammation and CRS test are suitable for in functional ELISA, and other immunoassays in diagnostics. The antibody can act as a capture antibody and detection antibody.

Classification: Astrovirus (Aastrovirus) human adenovirus anthrax bacillus acute respiratory infectious diseases Jejunum campylobacter candida Cytomegalovirus (CMV) Cryptosporidium (Crypto) parvum EB virus ebola virus (EV) Food source pathogenic bacteria resistant FluA FluB H. pylori Hepatitis b virus (HBV) HCV HIV1 HIV2 Human papilloma virus (HPV) herpes simplex virus HSV-1 HSV-2 Listeria monocytogenes Mycobacterium tuberculosis malaria Norovirus (NV, also called Winter Vomiting Bug) Prion Rift Valley Fever (RVF) rotavirus rubella virus salmonella Sexually transmitted diseases (STDs) TP(Treponema Pallidum) toxoplasma Vaccinia virus (cowpox virus) Vibrio cholerae West Nile virus (WNV)

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