Fungi and parasites

Definitive diagnosis of fungal infections depends on a positive blood culture or histopathological evidence of deep-tissue invasion but the sensitivity of the blood culture, especially in invasive aspergillosis, is very low. Culture techniques are also time-consuming, taking up to several weeks - an unacceptable time for the initiation of specific treatment of fungal infection. Alternative techniques to the traditional methods, such as enzyme linked immunosorbent assay detection of the...

Bacterial detection and quantification

Rapid detection by real-time PCR has been an important advantage where early diagnosis and appropriate antibiotic therapy are vital for survival, and traditional methods are often time-consuming. For instance, the early detection of bacterial DNA in the blood of critically ill patients with traditional culture diagnostic is still technically difficult (Cursons et al., 1999). Real-time PCR assays have been developed for quantification of different bacteria, including Chlamydia pneumoniae in...

Detection chemistries used in realtime PCR

There are three basic methodologies commonly used in the detection of RNA or DNA targets by real-time PCR and all of them utilize fluorescent dyes. In each case, a low initial fluorescent signal is increased proportionally during each succeeding PCR cycle in tandem with the exponential increase in the DNA product(s) formed. The simplest assay system involves the incorporation of a free dye into the newly formed double-stranded DNA product. The most used dye for this purpose in real-time PCR is...

Realtime PCR instrumentation an overview

It would be impractical to present technical information for every real-time instrument on the market today. However, it is important that users of realtime instruments have a basic understanding of how their instruments work and have some understanding of their physical capabilities and limitations. There are three major components in any real-time instrument 1) the light source, which determines the range of reporter dyes the instrument is capable of using 2) the detection system, where the...

Protocol 171 Cleaning chorionic villi CVS by microscopic dissection

The CVS sample is usually sent from the gynecology clinic in a universal vial in 10-15 ml of saline or culture medium. Sometimes small pieces of maternal tissue are co-biopsied with the villi, and other times a considerable amount of maternal blood is also present in the vial along with the CVS. The villi have a distinct, white frond-like appearance, as opposed to the reddish-pink amorphous appearance of other tissues, which in most cases can be distinguished visually. It is essential that any...