圓二色譜circulardichroism(cd)(文件)
【正文】 Determination of Protein Concentration 精確的方法有： 1 定量氨基酸分析； 2 用縮二脲方法測量多肽骨架濃度 或測氮元素的濃度 ； 3 在完全變性條件下測芳香氨基酸殘基的吸收，來確定蛋白質(zhì)的準(zhǔn)確濃度 . ? Not Acceptable: 1. Bradford Method. 2. Lowry Method. 3. Absorbance at 280 and/or 260 nm. Nitrogen flushing Flushing the optics with dry nitrogen is a must: ?Xe lamp has a quartz envelope, so if operated in air it’ll develop a lot of ozone, harmful for the mirrors ?below 195nm oxygen will absorb radiation HT plot ? The HT plot is very important, since readings above 600650V mean that not enough light is reaching the detector so a sample dilution or the use of shorter path cell are required. ? Furthermore the HT plot is in realty a single beam spectra of our sample, since there is a direct relation between HT and sample absorbance. By data manipulation HT conversion into absorbance and buffer baseline subtraction is possible. Alternatively single beam absorbance scale can be used already in CH2 during data collection, loosing however a bit the alerting functions of this channel. Bandwidth (SBW) selection ? Setting of slits should be as large as possible (to decrease noise level), but patible to the natural bandwidth (NBW) of the bands to be scanned. ? As a rule SBW should be kept at least 1/10 of the NBW, otherwise the band will be distorted. ? If NBW is not known a series of fast survey spectra at different SBW will help proper selection. Trade in of accuracy versus sensitivity (. the use of larger than theoretical SBW) is occasionally required. ? 2 nm in the far UV region ? 1 nm in the aromatic region (where fine structures may be present), optimal bandpass (as large as possible, but not loosing information) can be determined after a trial Number of data point ?data pitch, . number of data points per nm, will not directly influen
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鄂ICP備17016276號-1