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Genetic Engineering & Biotechnology News | GENengnews.com | JULY 2016 | 17 tion with isopropanol, and methanol were performed to determine extractable profle. Whether extracting with chloroform or isopropanol or evaluating via GC-MS or LC- MS, AntiBIND polypropylene (PP) plates consistently showed superior extraction performance when compared to fve other competitive grades of treated and untreated PP plates and specialty plates. Additionally, low protein binding plates offered in the market leached the hydrophilic moiety in the presence of these solvents exposing the hy- drophobic PP areas. (Check www.wheaton. com/AntiBIND for detailed experiments on extractable and leachable.) Using Advanced Plasma Technology to Alter the PP Properties from Hydrophobic to Hydrophilic to Minimize Protein-Wall Binding without the Addition of Leachables Polypropylene is a hydrophobic material made up of the non-polar monomer contain- ing CH, CH 2 , and CH 3 chemical groups. Most studies have shown that the adsorption process, or non-specifc protein binding, is reduced by decreasing the hydrophobicity of the sorbent surface. While this can be done by different mechanisms such as applying a hydrophilic coating (PEG-modifed surfaces and siliconization for glass) or incorporating hydrophilic compounds, these treatments are unstable and have the tendency to easily leach into solution exposing the hydrophobic parts of the plate. This was the case with the lead- ing brand of low protein binding plates in the presence of commonly used organic solvents. AntiBIND employs the use of Advanced Plasma Technology. Plasma surface activa- tion is the process by which surface polymer functional groups are replaced with different atoms from ions in the plasma to increase surface energy. Surface exposure to ener- getic species breaks down the polymer at the surface, creating free radicals. Free radicals quickly react with the material itself because they are unstable, which allows for the form- ing of stable covalent bonds. Proprietary mixture of plasmas made up of oxygen and water vapor were applied to the surface of the polypropylene modifying the surface of polypropylene with predominantly hydrogen-bond-acceptor uncharged polar groups. This treatment results in the forma- tion of a variety of oxygen- and nitrogen- con- taining polar groups including C-O-C bonds, C=O and O-C=O, and C-N. The resultant is an increased surface hydrophilicity as mea- sured by the decrease in water contact angle from 82° for the untreated polypropylene mi- croplates to 60° for the AntiBIND microplate. Signifcant Reduction in Protein Loss Since microplates are used in a plethora of applications that may include 1000s of different proteins, we devised various experi- ments to best capture these different factors in our protein adsorption and protein recov- ery experiments. To measure the amount of protein ad- sorbed to the plate, we measured the protein recovery rates of different fuorescently tagged proteins after different incubation times in the plates and at varied concentrations. The vari- ous sets of experiments were performed on the AntiBIND microplate, the best in class low protein binding plates, and standard high-quality PP plates. The results show a signifcant decrease in the adsorption (non- specifc protein binding) and consequently a signifcation increase in protein recovery. The AntiBind plate clearly demonstrates a signif- cant competitive advantage over the standard and low protein binding microplates com- mercially available today, particularly when the application deals with low protein con- centrations. The nonspecifc proteins binding process happened immediately after the addi- tion of the proteins with most of the binding occurring within the frst 4hrs. This trend was seen in all of the proteins that were tested and was very apparent at low concentrations. Robustness of Use and Compatibility with Automation for Chromatography Applications The American National Standards Insti- tute, Inc. (ANSI) and the Society for Labo- ratory Automation and Screening (SLAS) have defned the dimensional standards of microplates to ensure they work with most automated systems. The AntiBIND family of plates, 96 square well (0.5mL, 1.0 mL and 2.0 mL) and the 384 well (120 µL), are de- signed and manufactured according to these guidelines enabling compatibility with all automated systems. Moreover, they are pro- duced in class 7 clean room under Dnase/ Rnase free environment. The conical bottom of the 0.5mL and the round bottom of the (1.0 mL and 2.0 mL) allows for easy pipette access and maximum sample recovery. Addi- tionally, the 96-well plates have a raised rim that forms a tight seal, when using a sealing mat, to minimize sample loss due to evapo- ration and cross-talks between wells. In conclusion, even though microplates manufactured by different companies may look the same, there are major differences among them. The purity of the PP resin will have a tremendous effect on the integrity of the sample and the bioanalytical results. It is important to use the right raw materials (ho- modimer PP resin manufactured via metal- locene catalysts) with low additives. Low protein binding coatings and treatments are not all the same, and while they may work at high protein concentrations and in the pres- ence of common buffers, AntiBIND offers the superior protein recovery by as much as 3 fold at low protein concentrations and in the presence of harsh solvents. 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