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Research on Detecting the IgG Bioactive Content in Infant Formula Milk Powder

Author: HouFangNi
Tutor: ZhangJiaCheng
School: Qingdao Agricultural University
Course: Agricultural Products Processing and Storage Engineering
Keywords: IgG Infant formula milk powder Active content Enzyme - linked immunosorbent assay High Performance Liquid Chromatography
CLC: TS252.7
Type: Master's thesis
Year: 2009
Downloads: 101
Quote: 0
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Abstract

The development of a new generation of infant formula milk powder should it have, and breast milk is the same as or similar to the physiological functions, not only in the macro composition and content, but also at the micro simulated on breast milk. Therefore, to achieve real breast milk of infant formula milk powder, very important protective nutrients. The protective immune actives closer to breast milk is infant formula milk powder, and represents the development trend of the new generation of infant formula milk powder, immunoglobulin G - IgG is important in these protective immune actives species. Infant formula milk powder in IgG concentration is low compared with other IgG-containing products, it is still a lack of accurate and reliable measurement methods. Accordingly, this paper studies the detection of infant formula milk powder IgG activity content by ELISA and ELISA kit, ELISA kits and homemade HPLC method. The study concluded that for the detection of IgG in infant formula milk powder content of low activity parameters, and the eventual establishment of the determination of infant formula milk powder the IgG activity content - double antibody sandwich method. The main results are as follows: (1) standard bovine IgG activity content detection method: 5μg/mL rabbit anti-bovine IgG concentration stored at 37 ℃ 3h 4 ℃ 12h (overnight) coated microtiter plates; adopt containing 0.01g/mL blocking solution of BSA (bovine serum albumin) at 37 ℃ for 0.5h; optimum reaction time of 1.5h in the sample or standard bovine IgG; 1:250 times than diluted HRP-rabbit anti-bovine IgG as HRP antibody reaction 1h; substrate solution: the 50μL of TMB liquid substrate buffer 30μLH2O2 10mL, reaction 20min. The standard curve equation: y = 225.45x2 - 21.237x - 1.4713, correlation coefficient 0.999, the correlation of the standard curve. (2) by selecting the time of centrifugation and the solvent, experiment established the pretreatment conditions of the milk samples: The milk samples were dissolved in PBS solution to adjust the pH to between 4.5 to 4.6, 4000r/min centrifugation after 15min, filtered and taken supernatant , re-adjust the pH to 7.4, a constant volume in a 100-mL volumetric flask. This pretreatment method is simple and good separation. Double antibody sandwich (3) of this study is to establish the recoveries of 93.06% to 100.12% coefficient of variation of up to 3.090%, 1.200%, minimum; board and board between the coefficient of variation of the test results were less than 5% by SPSS software to make quality control charts can be learned, good stability of the method, the measured data with high accuracy. (4) the assembly of the kit stored at 4 ℃ can be saved 60d; learned that quality control charts can be made in accordance with the SPSS software, the stability of the kit, the measured data is accurate and reliable. Homemade kit of this study is to establish the test time savings largely (microtiter plates coated that do not need to save the 12h overnight, can be directly measured) to simplify the experimental procedure. HPLC method thereto compared to the obtained data is on the high side, low accuracy.

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