pna lectin  (Vector Laboratories)

Journal: bioRxiv

Article Title: Siglec-engaging immunosuppressive sialoglycans are upregulated in prostate cancer and are targetable to suppress bone metastasis

doi: 10.1101/2025.11.12.687981

Figure Lengend Snippet: ( A ) Detection of α2-3 linked sialylated glycans in CWR22Rv1 cells using α2-3 Lectenz flow cytometry. CWR22Rv1 cells were treated with a range of concentrations of E-612 for 24 hours. CWR22Rv1 treated with 1500nM E-612 had reduced levels of α2-3 Lectenz binding indicating a reduction in α2-3 linked sialylation in these cells. Reduced levels of α2-3 linked sialylated glycans were also detected in cells treated with E-612 using α2-3 Lectenz immunofluorescence. ( B ) Lectin flow cytometry and lectin immunofluorescence assays show CWR22Rv1 cells treated with E-612 for 24 hours have increased levels of binding to PNA lectin (which recognises galactosyl residues uncovered by the removal of sialic acids). ( C ) Detection of α2-3 linked sialylated glycans in mouse RM1 prostate cancer cells using α2-3 Lectenz flow cytometry. RM1 cells were treated with a range of concentrations of E-612 for 24 hours. RM1 cells treated with 1500nM E-612 had reduced levels of α2-3 Lectenz binding indicating a reduction in α2-3 linked sialylation in these cells. Reduced levels of α2-3 linked sialylated glycans were also detected in cells treated with E-612 using α2-3 Lectenz immunofluorescence. ( D ) Lectin flow cytometry and lectin immunofluorescence assays show RM1 cells treated with E-612 for 24 hours have increased levels of binding to PNA lectin (which recognises galactosyl residues uncovered by the removal of sialic acids). ( E ) HYDRA-7 flow cytometry shows treatment with 1500nM E-612 significantly reduces the binding of HYDRA-7 reagent to CWR22Rv1 cells indicating that E-612 has removed the sialoglycan ligands for Siglec-7 from the cell surface (unpaired t test, p=0.0207) ( F ) HYDRA-9 flow cytometry shows treatment with 1500nM E-612 significantly reduces the binding of HYDRA-9 reagent to CWR22Rv1 cells indicating that E-612 has removed the sialoglycan ligands for Siglec-9 from the cell surface (unpaired t test, p=0.0058).

Article Snippet: After another PBS wash, cells were blocked with 1X Carbo-FreeTM Blocking Solution (1X CFB) (Vector Laboratories, SP-5040-125) for 1 hr at room temperature, before they were incubated overnight in the dark at 4 °C in 1:1000 Fluorescein-labelled Peanut Agglutinin (PNA) Lectin (Vector Laboratories, FL-1071), 1:1000 Fluorescein-labelled Erythrina Cristagalli Lectin (ECL, ECA) (Vector Laboratories, FL-1141-5), 1:1000 Fluorescein-labelled Concanavalin A (Con A) Lectin (Vector Laboratories, FL-1001-25) or 1:400 SureLight® 488-conjugated SiaFind Alpha 2,3-Specific Lectenz® (Lectenz Bio, SK2301F).

Techniques: Flow Cytometry, Binding Assay, Immunofluorescence

Journal: bioRxiv

Article Title: Siglec-engaging immunosuppressive sialoglycans are upregulated in prostate cancer and are targetable to suppress bone metastasis

doi: 10.1101/2025.11.12.687981

Figure Lengend Snippet: ( A ) Dual immunofluorescence analysis of Siglec-E and CD14 (a myeloid marker) in PC3 prostate cancer tumours growing in mouse tibias. Siglec-E is co-localised with CD14 confirming its expression by myeloid cells within the murine prostate cancer TIME. Scale bar is 20□µm. ( B ) Using an RM1 intra-cardiac injection metastasis model, we investigated if therapeutic desialylation by E-612 can increase survival times of mice with metastatic prostate cancer. ( C ) Twice weekly dosing with 10mg/kg E-612 via intra-peritoneal injection prolongs the median survival rates of mice with metastatic prostate cancer by 53%. ( D ) Using an RM1 intra-caudal injection bone metastasis model we tested if E-612 mediated tumour desialylation can suppress the growth of bone metastatic prostate tumours. ( E ) Systemic treatment with E-612 (twice weekly dosing with 10mg/kg E-612 via intra-peritoneal injection) significantly reduces prostate cancer bone metastasis tumour burden (unpaired t test, p=0.0456). ( F ) PNA lectin flow cytometry analysis of white blood cells from mice treated with E-612. Systemic E-612 therapy significantly increases the levels PNA lectin binding to circulating immune cells (PNA lectin recognises galactosyl residues that are uncovered by the removal of sialic acids ) (Welch’s t test, p<0.0001).

Article Snippet: After another PBS wash, cells were blocked with 1X Carbo-FreeTM Blocking Solution (1X CFB) (Vector Laboratories, SP-5040-125) for 1 hr at room temperature, before they were incubated overnight in the dark at 4 °C in 1:1000 Fluorescein-labelled Peanut Agglutinin (PNA) Lectin (Vector Laboratories, FL-1071), 1:1000 Fluorescein-labelled Erythrina Cristagalli Lectin (ECL, ECA) (Vector Laboratories, FL-1141-5), 1:1000 Fluorescein-labelled Concanavalin A (Con A) Lectin (Vector Laboratories, FL-1001-25) or 1:400 SureLight® 488-conjugated SiaFind Alpha 2,3-Specific Lectenz® (Lectenz Bio, SK2301F).

Techniques: Immunofluorescence, Marker, Expressing, Injection, Flow Cytometry, Binding Assay