Establishment of Lectin-ELISA for the detection of multi-antennary AAG and its preliminary application

doi:10.3969/j.issn.1673-8640.2020.11.022

Abstract

Abstract:

Objective To establish a lectin enzyme-linked immunosorbent assay（Lectin-ELISA） for the detection of multi-antennary alpha 1-acid glycoprotein（AAG）,and to investigate the clinical application role of multi-antennary AAG in hepatocellular carcinoma（HCC）. Methods A Lectin-ELISA for detecting datura stramonium agglutinin（DSA）-AAG was established based on the principle that DSA can specifically identify the AAG of multi-antennary structure. This method was used to detect serum DSA-AAG levels of 220 HCC patients（HCC group）,237 disease controls [disease control group,74 cases of intrahepatic cholangiocarcinoma（ICC）（ICC group）,120 cases of liver cirrhosis（LC）（LC group）,43 cases of chronic hepatitis B（CHB）（CHB group）] and 80 healthy subjects（healthy control group）. Meanwhile,the pathological data and laboratory determination results of HCC patients were collected. Pearson correlation analysis was performed. Logistic regression method was used to establish the multi-index joint determination model. Receiver operating characteristic（ROC） curves were used to evaluate the efficiency of single and combined determinations in the diagnosis of HCC. Results The linear regression coefficient（r²） of serum DSA-AAG determined by Lectin-ELISA was 0.976,and the maximum between-run coefficient of variation（CV） was 10.51%. Free bilirubin,binding bilirubin,hemolysis and chylous had no effect on serum DSA-AAG determined by Lectin-ELISA. Serum DSA-AAG level of HCC group was higher than those of disease control group and healthy control group（P<0.001）,and serum AAG levels of HCC group and disease control group were higher than that of healthy control group（P<0.001）. Serum levels of DSA-AAG and AAG in cancer group（HCC+ICC） were higher than those in non-cancer group（LC+CHB+healthy control）（P<0.001）. Serum DSA-AAG level of HCC group was higher than that of ICC group（P<0.001）,while serum AAG level was lower than that of ICC group（P<0.001）. There was a negative correlation between DSA-AAG and alpha-fetoprotein （AFP） in HCC group（r=-0.029,P<0.05）. There was no statistical significance in serum DSA-AAG level among HCC patients with different tumor sizes and stages（P>0.05）. The areas under curves（AUC） of DSA-AAG,AAG and AFP single and combined determination models Logit_AAG1 for differentiating HCC from non-HCC were 0.651,0.632,0.803 and 0.836,respectively. The AUC of DSA-AAG and AAG single and combined determination model Logit_AAG2 for the differential diagnosis of AFP negative HCC and non-HCC were 0.669,0.607 and 0.929,respectively. Conclusions DSA-AAG and combined determination model may play roles in the differential diagnosis of HCC and AFP negative HCC.

Key words: Alpha 1-acid glycoprotein, Glycosylation, Lectin, Hepatocellular carcinoma

CLC Number:

R446.1

GUAN Wenqian, GAO Zhiyuan, FENG Huijuan, HONG Song, HE Yutong, HE Lu, GAO Chunfang. Establishment of Lectin-ELISA for the detection of multi-antennary AAG and its preliminary application[J]. Laboratory Medicine, 2020, 35(11): 1177-1185.

Figures/Tables 12

References 28

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组别	例数	AST/（U/L）		ALT/（U/L）		GGT/（U/L）		TB/（μmol/L）
HCC组	220	29.00（20.00~42.00）		29.00（20.00~42.00）		56.00（34.00~104.5）		14.00（10.90~18.38）
CHB组	43	223.00（88.00~519.00）		295.00（183.00~767.00）		114.00（63.00~165.00）		23.40（14.70~69.10）
LC组	120	32.00（24.25~44.75）		27.00（18.00~40.25）		46.50（28.25~79.75）		23.85（15.00~40.15）
ICC组	74	23.00（18.00~35.00）		24.50（16.00~34.25）		84.50（61.75~113.00）		14.35（10.70~18.43）
正常对照组	80			20.00（16.00~25.75）				14.65（12.00~16.63）
统计值		17.295		17.744		2.271		18.798
P值		<0.001		<0.001		0.132		<0.001
组别	DBil/（μmol/L）		Alb/（g/L）		TP/（g/L）		AFP/（μg/L）
HCC组	5.50（4.10~7.10）		42.01±3.74		69.62±8.28		61.00（4.70~1 210.00）
CHB组	9.60（5.20~47.00）		37.69±4.24		69.09±6.26		8.31（3.71~45.47）
LC组	10.20（6.00~20.00）		33.08±10.18		63.63±16.81		3.60（2.23~8.84）
ICC组	5.15（3.85~6.80）		42.26±3.46		68.67±5.12		2.90（2.00~4.70）
正常对照组			47.91±1.97		75.87±3.20		3.50（2.60~3.50）
统计值	22.056		46.845		161.700		82.406
P值	<0.001		<0.001		<0.001		<0.001

组别	例数	AST/（U/L）		ALT/（U/L）		GGT/（U/L）		TB/（μmol/L）
HCC组	220	29.00（20.00~42.00）		29.00（20.00~42.00）		56.00（34.00~104.5）		14.00（10.90~18.38）
CHB组	43	223.00（88.00~519.00）		295.00（183.00~767.00）		114.00（63.00~165.00）		23.40（14.70~69.10）
LC组	120	32.00（24.25~44.75）		27.00（18.00~40.25）		46.50（28.25~79.75）		23.85（15.00~40.15）
ICC组	74	23.00（18.00~35.00）		24.50（16.00~34.25）		84.50（61.75~113.00）		14.35（10.70~18.43）
正常对照组	80			20.00（16.00~25.75）				14.65（12.00~16.63）
统计值		17.295		17.744		2.271		18.798
P值		<0.001		<0.001		0.132		<0.001
组别	DBil/（μmol/L）		Alb/（g/L）		TP/（g/L）		AFP/（μg/L）
HCC组	5.50（4.10~7.10）		42.01±3.74		69.62±8.28		61.00（4.70~1 210.00）
CHB组	9.60（5.20~47.00）		37.69±4.24		69.09±6.26		8.31（3.71~45.47）
LC组	10.20（6.00~20.00）		33.08±10.18		63.63±16.81		3.60（2.23~8.84）
ICC组	5.15（3.85~6.80）		42.26±3.46		68.67±5.12		2.90（2.00~4.70）
正常对照组			47.91±1.97		75.87±3.20		3.50（2.60~3.50）
统计值	22.056		46.845		161.700		82.406
P值	<0.001		<0.001		<0.001		<0.001

A值	x	s	CV/%
高	0.867	0.08	9.23
中	0.730	0.07	9.59
低	0.666	0.07	10.51

A值	x	s	CV/%
高	0.867	0.08	9.23
中	0.730	0.07	9.59
低	0.666	0.07	10.51

组别	DSA-AAG（A值）	AAG/（g/L）
HCC组	0.47±0.07^*#	0.63±0.30^*
疾病对照组	0.44±0.09^*	0.66±0.36^*
正常对照组	0.39±0.10	0.45±0.15