title: Development and validation of a sensitive, high-throughput bioassay for the adhesion of radiolabeled E. coli to uroepithelial cells in vitro.
abstract: Vaccinium macrocarpon (cranberry) products have been used to prevent uropathogenic Escherichia (E.) coli adherence to uroepithelial cells (UEC) and may help reduce risk of urinary tract infection. Reported herein are the development and validation of an assay to assess antiadhesion activity of V. macrocarpon extracts and human urine. P-fimbriated E. coli (CFT073) was labeled with ³H-uridine, then co-incubated with HTB-4 UEC at a 400:1 ratio. V. macrocarpon extracts (0-17 mg proanthocyanidins/mL) were added to ³H-labeled E. coli before co-incubating with UEC. The assay yielded a sensitive inhibition curve: the lower limit of detection and half-maximal inhibitory concentration were 0.43 and 1.59 mg proanthocyanidins/mL for V. macrocarpon extract CEP 55; intra- and interassay coefficients of variance were <10% and <15%, respectively. V. macrocarpon extract CEP 3283 showed identical adhesion inhibition. Serial dilutions of urine from human participants who consumed V. macrocarpon beverages showed a linear decrease in antiadhesion activity. Antiadhesion assays conducted with urine from a human intervention study also showed good agreement with results obtained using the hemagglutination assay. Therefore, a sensitive, high-throughput, biologically relevant antiadhesion assay using ³H-E. coli co-incubated with UEC is reported, which can be used for studying the action of V. macrocarpon bioactives.
authors: School of Food Science, Washington State University , Pullman, Washington 99164-6376, United States.
authors: Bridget D
authors: BD
authors: Mathison
authors: Lindsey L
authors: LL
authors: Kimble
authors: Kerrie L
authors: KL
authors: Kaspar
authors: Christina
authors: C
authors: Khoo
authors: Boon P
authors: BP
authors: Chew
mesh_terms: D001422:Bacterial Adhesion / Q000187:drug effects; D001681:Biological Assay / Q000379:methods*; D006801:Humans; D044945:Proanthocyanidins / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology*; D014552:Urinary Tract Infections / Q000517:prevention & control* / Q000652:urine; D056689:Uropathogenic Escherichia coli / Q000187:drug effects*; D019459:Urothelium / Q000187:drug effects; D029799:Vaccinium macrocarpon / Q000737:chemistry*
chemical_list: D044945:Proanthocyanidins