Millex-GV filter

The reaction mixture was centrifuged (1450 X g), and the resultant clear supernatant was loaded onto a Cig column (5 g, 125 A) that had been pretreated with MeOH and conditioned with water (30 mL) prior to the loading of the centrifuged sample. The colurrm was washed with water (100 mL) then 1% acetonitrile (70 mL). The desired trisaccharide product was eluted with MeOH (HPLC grade, 120 mL). This eluate was concentrated under reduced pressure, and the product was dissolved in water (10 mL). This solution was passed through a Millex-GV filter (Millipore) to remove any residues of Cig resin. The filtrate was lyophilized to yield product as a fluffy white powder (133 mg, 196 mmol, 82%). NMR spectroscopy confirmed that the conversion was complete. 

M NaCl Dissolve 175 mg NaCl in 1 ml of water. Filter-sterilize through a 0.22-fxm Millex-GV filter unit using a 1-ml syringe. 

The fibronectin solution was filtered using Millex-GV filters (0.22 mm, Millipore", USA) and then divided in 1 ml portions for storage. Samples of pure fibronectin were stored at -20°C. Fibronectin concentration in the samples varied from 0.5 to 1 mg/ml. Fibronectin concentration in the samples was determined spectrophotometrically taking into account that 1.0% fibronectin solution absorbs 12.8 optical units at 280 nm. 

Further purification of X was initially thwarted by its disappearance, eventually attributed to its binding to the Millex-GV filter used for concentration between chromatographic steps [19]. Subsequent work led to amino terminal sequence data and the observations that X was very cross reactive with... 

To sterilize the RNase fusion proteins, use Millipore Millex-HV (Millipore Products Division, Bedford, MA) filters. Millex-GV low protein binding filters (Millipore Products) have been tried and result in a substantial loss of protein. 

Protein Purification. Canine plasma fibronectin was used in this study in order to correlate these in vitro studies with canine ex vivo experiments involving preadsorbed canine proteins. Canine FN was isolated from citrated canine plasma using the methods of Ruoslahti (11). The FN was suspended in phosphate buffered saline (PBS) containing 0.02% NaN3, and then snapfrozen and stored at -70 C until less than 24 hours before use. The protein was then snapthawed at 40 C, filtered (0.22 ym Millex GV, Millipore,... 

Petri dishes (140 mm. Cat. No. 501V) were obtained from Bibby Sterilin, and 0.2-//m filters (Falcon 7107) and replica dishes (Falcon 3047) fromBecton Dickinson. Millex-GV 0.22-//m filter units (Cat. No. SLGV 025 BS) were purchased from Millipore, 1-ml syringes from Terumo Medical Europe N.V., and 1-ml cuvettes (Cat. No. 1938 PS) from Kartell. 

Preparation of Amphiphile Solutions, (a) A quantity (15.0 mg) of DDP04(NH4)2 was dissolved in 5 mL of high-purity water by heating to ca. 50 °C, and the volume was subsequently adjusted to 100 mL with water, (b) A quantity (15.8 mg) of OH-DDPO4-(NH4)2 was dissolved in 5 mL of high-purity water by heating to ca. 80 °C. The solution was cooled to room temperature, filtered through a 0.22 pm filter (MILLEX-GV, MILLIPORE, Bedford, MA), and adjusted to a volume of 100 mL. 

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