A common rate-limiting step in many high throughput proteome scale analyses is the cloning of predicted open reading frames (ORFs) into technique-specific vectors. For example, methodologies for the detection of protein interactions such as Yeast-two-hybrid (Y2H) assay require validation using alternative methods as Protein Fragment Complementation Assays (PCA) or pulldown experiments. Various ...

Methods A case of oral allergy syndrome to sapodilla and custard apple was investigated following approval by Institutional Ethics Committee. Sapodilla allergy was confirmed by diagnostic tests (SPT and allergen-specific IgE). Sapodilla proteins were separated on SP-Sepharose by adsorption at pH 4 followed by step elution at pH 5 (SP1), and with increasing NaCl – 0.1 M (SP2) and 0.2 M (SP3). Fo...

Detection of vector-borne pathogens is necessary for investigation of their association with vertebrate and invertebrate hosts. The ability to detect Ehrlichia spp. within individual experimentally infected ticks would be valuable for studies to evaluate the relative competence of different vector species and transmission scenarios. The purpose of this study was to develop a sensitive PCR assay...

Quantitative PCR can often be improved by conducting the amplification with nested primers. First, fewer nonspecific amplification products, which could otherwise interfere with quantitation, are produced. Often, nonspecific products can be eliminated. In these cases, relatively simple nonspecific detection techniques are suitable for quantitation. In addition, nested primer PCR provides intrin...