Fast Multiplex Polymerase Chain Reaction on Boiled Clinical Samples for Rapid Diagnosis of Viral Infections
In spite of the recent dramatic development of new, rapid methods for detection of viral infections, most laboratory diagnosis of viral disease still depends either on the growth of virus or on the detection of a specific serologic reaction to infection. As tissue culture is the most widely sensitive and flexible, it remains the golden standard. But, because no single monolayer type is sensitive to the growth of all viruses most laboratories choose an array of different tissue cultures and, depending on the suspected viruses, inoculate each specimen into those lines or strains most likely to give a positive result. For the most part each virus, or virus group, produces a characteristic cytopathic effect, and the skilled technologist can usually develop a tentative identification on the basis of cell tropism and morphology, giving the clinician useful information at the earliest possible moment. Final identification of virus isolates usually depends on serologic tests and sometimes on morphologic, biophysical, or biologic characterization. The polymerase chain reaction (PCR) has the potential to replace these laborious viral isolation methods, but problems such as specificity, reproducibility, rapidity, cost, and the need for a common sequence in all strains of a particular virus must be taken into account. The production of low and high molecular weight non-specific PCR products must be noted while using most published PCR protocols. The accumulation of such PCR-products depletes primers and deoxynucleotide-triphosphates from the reaction mixture and leads to competition for enzyme with the desired PCR-product. Diagnosis must then rely on final detection assays such as hybridization with radioactive oligonucleotide probes to guarantee sufficient specificity, sensitivity and reproducibility.
KeywordsPolymerase Chain Reaction Protocol Polymerase Chain Reaction Procedure Long Control Region Cervical Scrape Polymerase Chain Reaction Yield
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