Page 12 - 麻醉与监护论坛2015年第5期

Cover Thesis

group: ISS, 32 cases of 25 or higher. Control group selected centrifugal 10min, 200ul suspension, 300ul biotin-
30 cases healthy elderly form hospital physical examination sheep anti mouse IgG antibody, 37℃ 30min, 3000rpm/
center. min, 10 min. 300ul SA-QD, 37℃ 30min, PBS washing
twice, 400ul suspension, FACSort flow cytometry
1.2 Test Methods instrument counting 3500 microspheres, take the
After 1 d, 3 d, 5 d, 7 d, 10 d, to extraction venous blood, FSC/SSC histogram, a scatter diagram and the FL-1
to separation serum, trauma each elderly patient extracted streaming histogram display microsphere surface average
20ul into mix serum (for the building method), control fluorescence intensity (MFI), X fluorescence intensity, Y
group each man also extracted 20ul into healthy mix serum number of microspheres.
(for the building method), and the method of quantitative
detection of PCT was measured. 1.4 Methodology Evaluation
To evaluate the sensitivity, linear range, precision and
1.2.1 Main Reagent positive point of this method.
Sheep anti human PCT polyclonal antibody, rat
anti human monoclonal antibody ( Ya Ji Biological 1.5 Preliminary Clinical Application
Technolog y Co., LTD. Shang hai, CHINA), 525 nm To extraction serum of High ISS group, low ISS group
quantum dots kit (including biotin sheep mouse IgG and control group, detected the serum PCT level by the self-
antibody, chain mildew resistance avidin quantum dots), created method, special protein analyzer detected CRP levels.
microsphere coupling agent (Bangs Laboratories, Inc.,
laboratory .USA), 10 um polystyrene microspheres 1.6 Statistical Analysis
(Tianjin university institute of materials, Tianjin, The SPSS17.0 statistical analysis was carried out on
CHINA), PBS: 0.01 mol/L, PH = 7.4. the test results, the measurement data compared by t test.

1.2.2 Main Instruments Fig.1 The SEM of PS microspheres
FACSort flow cytometry (Beckman Company, USA),
high-speed centrifuge (Sigma), oscillating instrument
(Huali Hot Melt Adhesive Co., LTD. Yiwu, CHINA),
752uv spectrophotometer (Xinmao Instrument Company,
Shanghai, CHINA), electronic microscope (OLYMPUS,
JAPAN).

1.3 Methodology Establishment Fig.2 The working curve of self- assay in PCT detection
To e x t r a c t i o n m i c ro s p h e re s s o l u t i o n , 1 0 0 0 g
centrifugal 10 min, 400ul Polylink suspension, 1000g
centrifugal 10min, 150ul PolyLink suspension, 10ul
EDC. HCL, blending, sheep anti human PCT antibody
200ul, 37℃ oscillation 1h, 1000g centrifugal 10 min,
the centrifugal PolyLink dilute into the appropriate
concentration (109-1012/L), store at 4℃. 200ul package
microspheres, adding serum specimen or the different
concentration of PCT (100ul), 37℃ oscillation 30min,
1000g centrifugal 10min, PBS washing, 1000g centrifugal
10min, 500ul suspension plus, 300ul rat anti PCT
monoclonal antibody, 37℃oscillation 30min, 1000g

Laboratory and Clinical ICnovveesrtigTahteiosins 93 FAM 2013 Mar/Apr Vol.20 Issue 2