Reverse lectin ELISA for detecting fucosylated forms of α1-acid glycoprotein associated with hepatocellular carcinoma

Abstract

Altered fucosylation of glycoproteins is associated with development of hepatocellular carcinoma (HCC). Lectins have been commonly used to assay changes in fucosylation of plasma glycoproteins. In the present study a recombinantly engineered form of the fucose binding lectin Aleuria aurantia (AAL) consisting of a single binding site for fucose (S2), was used to construct a reverse lectin ELISA method. Microtiter plates coated with the S2 lectin were used to capture glycoproteins from plasma samples followed by antibody detection of S2-bound fucosylated α1-acid glycoprotein (S2-bound AGP). The method was used to compare the level of S2-bound AGP in serum samples from a small cohort of patients with hepatitis, cirrhosis or HCC. Using the reverse S2 lectin ELISA it was shown that the levels of S2-bound AGP was significantly higher in HCC patients compared to non-cancer patients and that there was also a significant elevation of S2-bound AGP in HCC patients compared to cirrhosis patients. There was no correlation between the level of S2-bound AGP and total AGP concentration. The performance of S2-bound AGP in differentiating HCC from cirrhosis samples or hepatitis samples were compared to other markers. A combination of S2-bound AGP, α-fetoprotein and AGP concentration showed performances giving area under receiver operating curves of 0.87 and 0.95 respectively.

The cyanobacterial lectin, microvirin-N, enhances the specificity and sensitivity of lipoarabinomannan-based TB diagnostic tests

“…The sub-picomolar binding affinity of MVN to ManLAM and opportunistic bioconjugation framework made it an ideal capture reagent, while the ubiquitous use of antibodies as detection conjugates in ELISAs prompted the selection of Ab28 as the detection conjugate. 34,35 The capture bead was generated by loading streptavidin magnetic Dynabeads with biotinylated MVN. 21 An "all-in-one" OB-ELISA format was employed because it shortened overall assay time by 30+ minutes when compared to sequential addition without a significant impact on the signal-to-noise ratio (Fig.…”

Evaluation of AGP Fucosylation as a Marker for Hepatocellular Carcinoma of Three Different Etiologies

“…Serum AGP is synthesized mainly by hepatocytes in the liver, with approximately 45% of molecular weight contributed by N -glycans 14 . Changes in serum AGP glycosylation are highly correlated with HCC progression 19–21 ; however, it is unclear how N -glycan structures of AGP alter from liver cirrhosis to early HCC in different etiologies. In this study, we performed a comprehensive comparison of N -glycan structures of serum AGP between HCC and cirrhosis patients of the three common etiologies (i.e.…”

“…Cancer-associated glycosylation aberrations provide a rationale for discovering new biomarkers using glycomic/glycoproteomic technologies 23–28 . Previous studies revealed that although there was an increase in fucosylation level in both cirrhosis and HCC patients, HCC patients expressed AGP with more multifucosylated glycans 19,20 . Zhang et al .…”

“…Chronic liver disease and cirrhosis have been shown to affect the expression of AGP 18 and its glycosylation changes in both branching degree and fucosylation degree. Aberrant glycosylation of AGP is highly related to the carcinoma progression of HCC 19–21 , especially with elevated fucosylation 22 . However, it is still not clear how AGP glycoforms alter during the progression from liver cirrhosis to HCC of different etiologies.…”

The fucose‐specific lectin ANL from Aspergillus niger possesses anti‐cancer activity by inducing the intrinsic apoptosis pathway in hepatocellular and colon cancer cells

“…A well‐known fucose‐binding lectin from the Orange peel fungus Aleuria aurantia (AAL), recently bioengineered as a monomeric form AAL‐S2 with high affinity towards α1–6 fucosylated (core‐fucosylated) glycans 32 was used to construct a reverse‐lectin ELISA method to support identification of aberrantly fucosylated alpha‐1 acid glycoprotein in the serum of HCC patients 33 . Immuno‐histochemical studies revealed its diagnostic potential by differentially binding to normal and metastatic cancerous tissues 34 .…”