continuous–elution electrophoresis mini prep cell assembly system  (Bio-Rad)

Journal: International Journal for Parasitology

Article Title: Excreted/secreted Schistosoma mansoni venom allergen-like 9 (SmVAL9) modulates host extracellular matrix remodelling gene expression

doi: 10.1016/j.ijpara.2014.04.002

Figure Lengend Snippet: Schistosoma mansoni venom allergen-like 9 (SmVAL9) undergoes transcriptional regulation throughout schistosome development, is maximally expressed in miracidia and is post-translationally modified in eggs. (A) DNA microarray analysis of SmVAL9 expression throughout 15 lifecycle stages. Histogram represents normalised mean fluorescent intensities + S.D. ( n = 3 replicates/lifecycle stage except adult female where n = 2) of SmVAL9 transcript abundance derived from oligonucleotide CD111828 as described previously . Dashed box encloses schistosome lifecycle stages where SmVAL9 is maximally expressed. Inset drawing represents SmVAL9 (Smp_176180) gene organisation (five exons – black boxes; four introns – black lines) and localisation of oligonucleotide CD111828 to exon 5 (SchistoGeneDB v5.1). (B) Expression and purification of recombinant SmVAL9 (rSmVAL9) in Escherichia coli cells. The majority of rSmVAL9 is found in insoluble inclusion bodies after induction, allowing preparatory cell electrophoresis (Prep. Cell) to concentrate and purify large amounts of the protein for murine immunisations and other downstream studies. (C) Anti-rSmVAL9 recognises the 19.3 kDa rSmVAL9 immunogen and a ∼25 kDa post-translationally modified, native SmVAL9 (nSmVAL9) from soluble egg antigen. Pre-bleed normal mouse serum does not recognise either protein or the BSA control.

Article Snippet: Purification of rSmVAL9 was carried out by size fractionation using a continuous elution electrophoresis apparatus (Model 491 Prep Cell; Bio-Rad, UK), following the manufacturer’s instructions.

Techniques: Modification, Microarray, Expressing, Derivative Assay, Purification, Recombinant, Electrophoresis, Control