Value of peripheral blood monocyte subsets and CD64 expression in the diagnosis and prognosis of sepsis / 中华危重病急救医学

Objective:To explore the value of monocyte subsets and CD64 expression in the diagnosis and prognosis of sepsis.Methods:A prospective case-control study was designed. 30 septic patients and 30 non-septic patients who were admitted to the intensive care unit (ICU) of the PLA Army Characteristic Medical Center from March 2021 to March 2022 were enrolled. After 1, 3, and 5 days of ICU admission, peripheral blood samples were taken from patients. Flow cytometry was used to detect the proportion of monocyte subsets and the expression level of CD64 on the surface, and the difference of expression between patients in two group was analyzed. The risk variables for sepsis were analyzed using single-factor and multi-factor Logistic regression. The diagnostic efficacy of each risk factor for sepsis was determined using the receiver operator characteristic curve (ROC curve).Results:One day after ICU admission, the proportions of monocytes and classic monocytes in white blood cells (WBC) of septic patients were significantly lower than those of non-septic patients [proportion of monocytes to WBC: (4.13±2.03)% vs. (6.53±3.90)%, proportion of classic monocytes to WBC: 1.97 (1.43, 2.83)% vs. 3.37 (1.71, 5.98)%, both P < 0.05]. The proportion of non-classical monocytes in monocytes was significantly higher in septic patients than that in non-septic patients [(11.42±9.19)% vs. (6.57±4.23)%, P < 0.05]. The levels of CD64 expression in monocytes, classic monocytes, intermediate monocytes and non-classic monocytes were significantly higher in sepsis patients than those in non-septic patients [mean fluorescence intensity (MFI): 13.10±6.01 vs. 9.84±2.83 for monocytes, 13.58±5.98 vs. 10.03±2.84 for classic monocytes, 13.48±6.35 vs. 10.22±2.99 for intermediate monocytes, 8.21±5.52 vs. 5.79±2.67 for non-classic monocytes, all P < 0.05]. Multivariate Logistic regression research showed that CD64 in typical monocytes [odds ratio ( OR) = 1.299, 95% confidence interval (95% CI) was 1.027-1.471, P = 0.025] and the proportion of non-typical monocytes in monocytes ( OR = 1.348, 95% CI was 1.034-1.758, P = 0.027) were the independent risk factors for sepsis. ROC curve showed that the area under the ROC curve (AUC) of CD64 expression of classical monocytes, the fraction of non-classical monocytes in monocytes, and procalcitonin (PCT) in the diagnosis of sepsis was 0.871. A correlation analysis revealed a negative relationship between the acute physiology and chronic health status evaluation Ⅱ (APACHE Ⅱ) on the first, third, and fifth days following ICU admission and the expression level of CD64 in patients' classic monocytes ( r values were -0.264, -0.428 and -0.368, respectively, all P < 0.05). Conclusions:Combining the proportion of non-classical monocytes in monocytes, the level of plasma PCT, and the CD64 expression of classic monocytes in peripheral blood has good efficacy in identifying sepsis and assessing its severity.

Prediction of maximum absorption activities with polyclonalantibodies immobilized on magnetic submicron particles torecognize positive mutants / 西安交通大学学报(医学版)

Objective To observe polyclonal antibodies immobilized on magnetic submicron particles (MSP) as affinity adsorbents and test the reliability of predicted maximum adsorption activity of an enzyme/mutant from a cell lysate (Vs) in recognizing positive mutants.Methods Escherichia coli alkaline phosphatase (ECAP) and Pseudomonas Aeruginosa arylsulfatase (PAAS) were purified by affinity chromatography to serve as immunogens for the preparation of their antisera, which after fractionation by 33％ ammonium sulfate and DEAE-cellulose chromatography yielded the respective polyclonal antibodies.After activation of COOH on MSP, polyclonal antibodies of each enzyme were immobilized to give MSP-polyAb.Activities of an adsorbed enzyme were measured with a chromogenic substrate of 4-nitrphenol by determining absorbance at 405nm after the termination of reaction by alkali.Based on the response curve of activities of the adsorbed enzyme to protein quantities of a lysate, Vs was predicted for comparison.Results The maximum adsorption quantity of ECAP or PAAS on the respective MSP-polyAb was about 2.0mg/g.Specific activity of ECAP after affinity purification was about 70-fold of that of PAAS.ECAP mutant R168K showing about 50％ activity improvement versus ECAP was recognized by comparison of Vs predicted with only 2.5μg of MSP-polyAb;with PAAS mutant G138S as the starting one, the use of 10.0μg of MSP-polyAb to predict Vs recognized the mutants bearing more than 20％ activity improvement.Conclusion With an optimized quantity of MSP-polyAb to predict Vs, weak positive mutants of an enzyme of low activity can be recognized when activities of the adsorbed enzyme/mutant are reliably measured.