Subbaraman Lakshman N, Glasier Mary Ann, Senchyna Michelle, Sheardown Heather, Jones Lyndon
Centre for Contact Lens Research, School of Optometry, University of Waterloo, Waterloo, Ontario, Canada.
Eye Contact Lens. 2007 Jul;33(4):169-73. doi: 10.1097/01.icl.0000248155.23800.20.
Extracting lysozyme from Food and Drug Administration group IV etafilcon lenses by using 0.2% trifluoroacetic acid and acetonitrile (TFA/ACN) is a well-established procedure. TFA/ACN has been the extraction buffer of choice for extracting proteins from silicone hydrogel contact lenses. The purpose of this study was to determine the efficiency of TFA/ACN in extracting lysozyme from silicone hydrogel and etafilcon lenses by using an in vitro model.
ACUVUE 2, Focus NIGHT & DAY, O2 Optix, PureVision, and ACUVUE Advance lenses were incubated in simple lysozyme solution and a complex artificial tear solution consisting of multiple tear components containing lysozyme labeled with iodine 125. All the silicone hydrogel lenses were incubated for 28 days, whereas the ACUVUE 2 lenses were incubated for 7 days at 37 degrees C with constant rotation. After the incubation period, radioactive counts were determined, and the lenses were placed in an appropriate volume of the buffer for 24 hours in darkness. The lenses were removed from the buffer, and radioactive counts were determined again.
Extraction efficiencies for lysozyme from the artificial tear solution were 97.2% +/- 1.2% for ACUVUE 2, 64.3% +/- 6.2% for Focus NIGHT & DAY, 62.5% +/- 5.6% for O2 Optix, 53.5% +/- 5.8% for PureVision, and 89.2% +/- 3.4% for ACUVUE Advance. Results were similar for the lysozyme extracted after incubating in the simple lysozyme solution.
TFA/ACN is extremely efficient at extracting lysozyme deposited on etafilcon lenses. However, it does not extract all the lysozyme deposited on silicone hydrogel lenses, and alternative extraction procedures should be sought.
采用0.2%三氟乙酸和乙腈(TFA/ACN)从食品药品监督管理局IV类依他氟酮镜片中提取溶菌酶是一种成熟的方法。TFA/ACN一直是从硅水凝胶隐形眼镜中提取蛋白质的首选提取缓冲液。本研究的目的是通过体外模型确定TFA/ACN从硅水凝胶和依他氟酮镜片中提取溶菌酶的效率。
将ACUVUE 2、焦点日夜型、O2 Optix、PureVision和ACUVUE Advance镜片分别置于简单溶菌酶溶液以及含有多种泪液成分且溶菌酶用碘125标记的复合人工泪液溶液中孵育。所有硅水凝胶镜片孵育28天,而ACUVUE 2镜片在37℃下持续旋转孵育7天。孵育期结束后,测定放射性计数,然后将镜片置于适量缓冲液中在黑暗中放置24小时。将镜片从缓冲液中取出,再次测定放射性计数。
从人工泪液溶液中提取溶菌酶的效率,ACUVUE 2为97.2%±1.2%,焦点日夜型为64.3%±6.2%,O2 Optix为62.5%±5.6%,PureVision为53.5%±5.8%,ACUVUE Advance为89.2%±3.4%。在简单溶菌酶溶液中孵育后提取的溶菌酶结果相似。
TFA/ACN在提取沉积在依他氟酮镜片上的溶菌酶方面极其有效。然而,它不能提取沉积在硅水凝胶镜片上的所有溶菌酶,应寻求替代提取方法。