gingivalis biofilm and how this relates to pathogeniCity. In our laboratory we have devised a reproducible

continuous culture method to grow biofilm and planktonic cells simultaneously in the same fermentor vessel. Using this approach we have compared the cell envelope proteome of P. gingivalis W50 biofilm and planktonic cells [15]. In this current study, we have PF-2341066 expanded our investigation of these cells, comparing the global gene expression within P. gingivalis biofilm and planktonic cells using microarray analysis. Methods Continuous culture conditions and biofilm formation The growth and physical characterization of the biofilm and planktonic cells analysed in this study have been described in Ang et al. [15]. The continuous culture system allows the simultaneous co-culture of planktonic cells and biofilm cells under identical growth conditions [15]. Briefly, the methods used were as follows. To produce biofilm and planktonic cells for RNA harvest P. gingivalis was grown in continuous culture, in duplicate, using a Bioflo

BAY 73-4506 chemical structure 110 fermentor with a total volume of 400 mL (New Brunswick Scientific, Edison, NJ, USA) in BHI medium supplemented with 5 mg mL-1 cysteine hydrochloride and 5.0 μg mL-1 haemin. Growth was initiated by inoculating the fermentor vessel with a 24 hour batch culture (100 mL) of P. gingivalis grown in the same medium. After a 24 h incubation the media reservoir pump was turned on and the media flow adjusted to give a dilution rate of 0.1 h-1(mean generation time of 6.9 h). The temperature of the vessel was maintained at 37°C and the pH at 7.4 ± 0.1. The culture was continuously gassed with 5% CO2 in 95% N2. Optical density readings (OD650 nm) and purity of the culture were analyzed daily. Biofilm could be seen to be forming on the fermentor vessel walls and on glass microscope slides that were fixed to the vessel walls. Each P. gingivalis W50 culture was maintained for 40 days until harvesting. Planktonic cells were harvested by rapidly pumping them out of the fermentor vessel. The microscope slides were then

removed from the fermentor vessel for examination of biofilm thickness and cell viability. The biofilm was rinsed twice with cold PGA buffer [16] to remove contaminating planktonic cells and then removed by scraping with a spatula and suspended in cold PGA FAD buffer in a 50 mL {Selleck Anti-diabetic Compound Library|Selleck Antidiabetic Compound Library|Selleck Anti-diabetic Compound Library|Selleck Antidiabetic Compound Library|Selleckchem Anti-diabetic Compound Library|Selleckchem Antidiabetic Compound Library|Selleckchem Anti-diabetic Compound Library|Selleckchem Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|Anti-diabetic Compound Library|Antidiabetic Compound Library|buy Anti-diabetic Compound Library|Anti-diabetic Compound Library ic50|Anti-diabetic Compound Library price|Anti-diabetic Compound Library cost|Anti-diabetic Compound Library solubility dmso|Anti-diabetic Compound Library purchase|Anti-diabetic Compound Library manufacturer|Anti-diabetic Compound Library research buy|Anti-diabetic Compound Library order|Anti-diabetic Compound Library mouse|Anti-diabetic Compound Library chemical structure|Anti-diabetic Compound Library mw|Anti-diabetic Compound Library molecular weight|Anti-diabetic Compound Library datasheet|Anti-diabetic Compound Library supplier|Anti-diabetic Compound Library in vitro|Anti-diabetic Compound Library cell line|Anti-diabetic Compound Library concentration|Anti-diabetic Compound Library nmr|Anti-diabetic Compound Library in vivo|Anti-diabetic Compound Library clinical trial|Anti-diabetic Compound Library cell assay|Anti-diabetic Compound Library screening|Anti-diabetic Compound Library high throughput|buy Antidiabetic Compound Library|Antidiabetic Compound Library ic50|Antidiabetic Compound Library price|Antidiabetic Compound Library cost|Antidiabetic Compound Library solubility dmso|Antidiabetic Compound Library purchase|Antidiabetic Compound Library manufacturer|Antidiabetic Compound Library research buy|Antidiabetic Compound Library order|Antidiabetic Compound Library chemical structure|Antidiabetic Compound Library datasheet|Antidiabetic Compound Library supplier|Antidiabetic Compound Library in vitro|Antidiabetic Compound Library cell line|Antidiabetic Compound Library concentration|Antidiabetic Compound Library clinical trial|Antidiabetic Compound Library cell assay|Antidiabetic Compound Library screening|Antidiabetic Compound Library high throughput|Anti-diabetic Compound high throughput screening| centrifuge tube. PGA buffer contained 10.0 mM NaH2PO4, 10.0 mM KCl, 2.0 mM citric acid, 1.25 mM MgCl2, 20.0 μM CaCl2, 25.0 μM ZnCl2, 50.0 μM MnCl2, 5.0 μM CuCl2, 10.0 μM CoCl2, 5.0 μM H3BO3, 0.1 μM Na2MoO4, 10 mM cysteine-HCl with the pH adjusted to 7.5 with 5 M NaOH. Biofilm characterization The viability of cells comprising the biofilms that were on the glass microscope slides were determined using LIVE/DEAD® BacLight™ stain as per manufacturer instructions (Invitrogen) with visualized using confocal laser scanning microscopy (CLSM) essentially as described by Loughlin et al. [17].