α 2 3 sialidase

Manufactured by Takara Bio

Sourced in China, Japan

α-2,3-sialidase is an enzyme that selectively cleaves α-2,3-linked sialic acid residues from glycoconjugates. It can be used for the analysis and characterization of glycan structures.

Automatically generated - may contain errors

Lab products found in correlation

Vibrio cholerae neuraminidase, by Roche (1 mentions) C18 ziptip pipette tip, by Merck Group (1 mentions) 4800 maldi tof tof analyzer, by Thermo Fisher Scientific (1 mentions) Sialidase a, by Agilent Technologies (1 mentions) Vibrio cholerae neuraminidase vcna, by Roche (1 mentions)

10 protocols using α 2 3 sialidase

Influenza Virus HA Assay Protocols

Check if the same lab product or an alternative is used in the 5 most similar protocols

The receptor-binding specificities of human influenza viruses were determined by
HA assays with 1% chicken and sheep red blood cell (cRBC and sRBC) suspensions.
For sialidase treatment, 90 μL of a 10% cRBC suspension was treated with 10 μL
of α-2,3-sialidase (50 mU/μL; TaKaRa, Dalian, China) for 10 minutes at 37°C. The
sample was then washed twice with PBS, centrifuged at 210 × gfor 5 minutes each time, adjusted to a final working concentration (1%) with
PBS, and stored at 4°C. For Vibrio cholerae neuraminidase
(VCNA; Roche, San Francisco, CA, USA) treatment, 90 μL of a 10% cRBC suspension
was treated with 10 μL of VCNA (50 mU/μL) for 1 hour at 37°C, washed twice with
PBS, centrifuged at 210 × g for 5 minutes each time, adjusted
to a final working concentration (1%) with PBS, and stored at 4°C. For the HA
assay, viruses were serially diluted 2-fold with 50 μL of PBS and mixed with
50 μL of a 1% RBC suspension in a 96-well plate. HA titers were read after 20
minutes of reaction at room temperature.

Zhang C., Cui H., Wang Z., Dong S., Zhang C., Li J., Meng K., Sun Y., Liu J., Guo Z, & Chen L. (2021). Pathogenicity and transmissibility assessment of two strains of human influenza virus isolated in China in 2018. The Journal of International Medical Research, 49(1), 0300060520982832.

+ Open protocol

+ Expand

Determining Viral Sialic Acid Receptor Specificity

Check if the same lab product or an alternative is used in the 5 most similar protocols

The receptor-binding specificity of the virus to α2,6-linked sialic acid receptor was determined by comparing the hemagglutination titers between the native and α2,3-linked-sialidase-treated cRBCs as described previously (27 (link)). PR8 virus was used as the control. Briefly, 3 ml of a 10% suspension of cRBCs was incubated with 0.5 U α2,3-sialidase (TaKaRa, Beijing, China) for 1.5 h at 37°C. After washing 3 times with PBS, the treated cRBCs were adjusted to a final concentration of 0.5% with PBS. The hemagglutination titer of each virus was determined with the treated cRBCs. The receptor-binding specificity of the virus to α2,3-linked sialic acid receptor was measured in a sialic acid competition assay. The disialoganglioside GD1a (GlycoSci, Shanghai, China), which contains α2,3-linked sialic acid receptor, was used to competitively bind virus. Briefly, allantoic fluid of each virus was first treated with 25 μM/ml zanamivir for 1 h at 37°C. Then, the virus was further incubated with 12.5 μg/ml GD1a overnight at 4°C. After incubation, the GD1a-treated virus and untreated virus were finally tested with normal 0.5% cRBCs.

Wang F., Wan Z., Wang Y., Wu J., Fu H., Gao W., Shao H., Qian K., Ye J, & Qin A. (2021). Identification of Hemagglutinin Mutations Caused by Neuraminidase Antibody Pressure. Microbiology Spectrum, 9(3), e01439-21.

+ Open protocol

+ Expand

Sialidase-treated Chicken Red Blood Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Sialidase-treated Chicken red blood cells (CRBCs) were prepared as previously described18 (link) with modified. CRBCs were washed and diluted to 10% (vol/vol) in PBS. 0.1 ml CRBCs were incubated with 10 μl (50 U/ml) of α2,3 –sialidase (Takara, Japan) for 15 min at 37 °C, washed three times in PBS, and re-suspended in PBS.

Sang X., Wang A., Ding J., Kong H., Gao X., Li L., Chai T., Li Y., Zhang K., Wang C., Wan Z., Huang G., Wang T., Feng N., Zheng X., Wang H., Zhao Y., Yang S., Qian J., Hu G., Gao Y, & Xia X. (2015). Adaptation of H9N2 AIV in guinea pigs enables efficient transmission by direct contact and inefficient transmission by respiratory droplets. Scientific Reports, 5, 15928.

+ Open protocol

+ Expand

Characterization of GM-CSF Glycosylation

Check if the same lab product or an alternative is used in the 5 most similar protocols

GM-CSF of the different cell lines (1-4 μg of protein in 20 μL) was supplemented with 10 μL of 3x Tricine gel loading buffer (1.5 M Tris-HCl, pH 8.45, 35% glycerol, 10% SDS, 0.01% coomassie and 30 mM of DTT) and incubated for 10 minutes at 98°C. 3 μL of a 500 mM iodoacetamide stock was added and the samples were incubated for 1 hour in the dark. We separated the samples on a 12% tricine SDS-PAGE gel and cut out the bands. Detailed methods for in-gel tryptic digestion are described in Supplementary Note 4. We treated the tryptic peptides with either no enzyme, 50 mU of α-2,3-sialidase (Takara Bio Inc.), or both 200 of mU A. ureafaciens sialidase and 2 mU of Streptococcus pneumoniae β-1,4-galactosidase (Prozyme). All digests were incubated for 24 hours at 37°C, dried in a speedvac, reconstituted with 10 μL of 0.2% trifluoroacetic acid (TFA) (Sigma-Aldrich) and cleaned up with C18 ZipTip^® pipette tips (Millipore) according to the manufacturer’s instructions. Samples were analysed with 6-aza-2-thiothymine (ATT) matrix saturated in 50% acetonitrile containing 0.1% TFA, on a 4800 MALDI TOF/TOF™ Analyzer (Applied Biosystems) in the positive ion mode. The reported m/z values were observed in several iterations of technical optimizations and the results of the fully optimized experiments are shown.

Meuris L., Santens F., Elson G., Festjens N., Boone M., Dos Santos A., Devos S., Rousseau F., Plets E., Houthuys E., Malinge P., Magistrelli G., Cons L., Chatel L., Devreese B, & Callewaert N. (2014). GlycoDelete technology: simplifying mammalian cell N-glycosylation for recombinant protein expression. Nature biotechnology, 32(5), 485-489.

+ Open protocol

+ Expand

Analyzing H5N6 Virus Receptor Binding

Check if the same lab product or an alternative is used in the 5 most similar protocols

The receptor-binding specificities of the three H5N6 viruses were determined by HA assays with 1% cRBC and sRBC suspensions. For sialidase treatment, 90 μL of a 10% cRBC suspension was treated with 10 μL of α-2,3-sialidase (50 mU/μL) (TaKaRa, Dalian, China) for 10 min at 37 °C. The sample was then washed two times with PBS, centrifuged at 1500 rpm for 5 min each time, adjusted to a final working concentration (1%) with PBS, and stored at 4 °C. For VCNA (Roche, San Francisco, CA) treatment, 90 μL of a 10% cRBC suspension was treated with 10 μL of VCNA (50 mU/μL) for 1 h at 37 °C, washed two times with PBS, centrifuged at 1500 rpm for 5 min each time, adjusted to a final working concentration (1%) with PBS, and stored at 4 °C. For the HA assay, viruses were serially diluted 2-fold with 50 μL of PBS and mixed with 50 μL of a 1% RBC suspension in a 96-well plate. HA titers were read after 20 minutes of the reaction at room temperature.

Zhao Z., Guo Z., Zhang C., Liu L., Chen L., Zhang C., Wang Z., Fu Y., Li J., Shao H., Luo Q., Qian J, & Liu L. (2017). Avian Influenza H5N6 Viruses Exhibit Differing Pathogenicities and Transmissibilities in Mammals. Scientific Reports, 7, 16280.

+ Open protocol

+ Expand

Glycan Analysis Protocol for Researchers

Check if the same lab product or an alternative is used in the 5 most similar protocols

The PA-glycans were purchased from Takara Bio Inc. (Kyoto, Japan), Seikagaku Corporation (Tokyo, Japan) or Masuda Chemical Industries Co., Ltd. (Takamatsu, Japan). The PA-glycans which were not commercially available were prepared as described elsewhere [5] (link), [7] . Reagents were purchased from Wako Pure Chemical Industries, Inc. (Osaka, Japan) or Nakalai Tesque, Inc. (Kyoto, Japan). Arthrobacter sialidase was purchased from Nakalai Tesque, Inc. and α2-3sialidase was from Takara Bio Inc.

Natsuka S., Masuda M., Sumiyoshi W, & Nakakita S.I. (2014). Improved Method for Drawing of a Glycan Map, and the First Page of Glycan Atlas, Which Is a Compilation of Glycan Maps for a Whole Organism. PLoS ONE, 9(7), e102219.

+ Open protocol

+ Expand

Exoglycosidase Treatment of PA-Glycans

Check if the same lab product or an alternative is used in the 5 most similar protocols

Exoglycosidase treatment of PA-glycans was carried out by using α2–3, –6-sialidase (C. perfringens: Merck), α2–3-sialidase (S. typhimurimum LT2, recombinant, E. coli: Takara Bio), β-galactosidase (Jack bean: ProZyme, Inc.), β-N-acetylhexosaminidase (Jack bean: ProZyme, Inc.), or α-L-fucosidase (bovine kidney: ProZyme, Inc.). PA-glycan was treated with the enzyme (50 mU) in 20 μl of 50 mM ammonium acetate buffer (β-galactosidase: pH 3.5, α2–3, −6-sialidase and β-N-acetylhexosaminidase: pH 5.0, α2–3-sialidase and α-L-fucosidase: pH 5.5), at 37 °C for appropriate periods (0.5 h for α2–3, –6-sialidase and α2–3-sialidase; 20 h for others). The enzyme reaction was terminated by heating at 99.9 °C for 5 min.

Hasehira K., Hirabayashi J, & Tateno H. (2016). Structural and quantitative evidence of α2–6-sialylated N-glycans as markers of the differentiation potential of human mesenchymal stem cells. Glycoconjugate Journal, 34(6), 797-806.

+ Open protocol

+ Expand

Sialic Acid Profiling with Lectin Array

Check if the same lab product or an alternative is used in the 5 most similar protocols

After Cy3-labeling, 20 μL of the reaction solution was digested into sialic acid with sialidase A (#GK80040; Agilent Technologies Inc.) or α2-3sialidase (#4455; Takara Bio Inc., Shiga, Japan), according to the manufacturer's instructions. The volume of the solution was adjusted to 100 μL with probing buffer and incubated at 4 °C until use. For analysis, 60 μL of the solution was applied to the lectin microarray. The data obtained for mice aged 2 and 23–25 months were compared with those without enzymatic reactions and the difference was calculated using the following formula: difference = log₂ (signal intensity with sialidase/signal intensity without sialidase).

Itakura Y., Hasegawa Y., Kikkawa Y., Murakami Y., Sugiura K., Nagai-Okatani C., Sasaki N., Umemura M., Takahashi Y., Kimura T., Kuno A., Ishiwata T, & Toyoda M. (2023). Spatiotemporal changes of tissue glycans depending on localization in cardiac aging. Regenerative Therapy, 22, 68-78.

+ Open protocol

+ Expand

Receptor-Binding Assay for Influenza Viruses

Cited 2 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

Receptor-binding property is a key determinant of the mammalian adaption of the influenza virus (Gao et al., 2009 (link); Zhang et al., 2012 (link)). Hemagglutination assays using resialylated chicken red blood cells (cRBCs) and sheep red blood cells (sRBCs) were performed to analyze the receptor-binding property of the CIVs. The surface of cRBCs expresses both human-like α2,6 and avian-like α2,3 sialic acid receptors (Ito et al., 1997 (link)), while sRBCs only contain avian-like α2,3 sialic acid receptors (Medeiros et al., 2001 (link)). The α2,3 sialic acids were removed from the cRBCs' surface by incubating with α2,3-sialidase (Takara, Dalian, Liaoning, China). The desialylation cRBCs were generated by incubating with Vibrio cholerae neuraminidase (VCNA, Roche, San Francisco, CA, United States). The desialylation cRBCs were used as the negative control. The A/Sichuan/1/2009(H1N1) and A/chicken/Hebei/3/2013(H5N2), which bind to α2,6-cRBCs and α2,3-cRBCs, respectively (Shi et al., 2017 (link)), were used as controls.

Meng B., Li H., Feng C., Guo W., Feng Y., Zhu D., Chen H, & Zhang Y. (2023). Emergence of a novel reassortant H3N6 canine influenza virus. Frontiers in Microbiology, 14, 1186869.

+ Open protocol

+ Expand

Influenza Virus Receptor Binding Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

Chicken red blood cells (CRBC, Beijing Farm Animal Research Center, Institute of Laboratory Animal Sciences, Chinese Academy of Medical Sciences) and α-2,3-sialidase (50 mU/μL, TaKaRa, Shiga, Japan) were mixed at a ratio of 9:1 and incubated at 37 °C for 10 min. After two washes with PBS, the solution was resuspended to create a 0.75% red blood cell (RBC) suspension. Next, 100 µL of 10% CRBC was mixed with 50 mU of Vibrio cholerae neuraminidase (VCNA, Sigma-Aldrich, St. Louis, MO, USA), incubated at 37 °C for 1 h, washed 3 times with PBS, and then mixed with 50 µL of PBS. CRBCs and sheep RBCs (TaKaRa, Japan) were prepared as 0.75% RBC suspensions to assess the agglutination characteristics of Th005 and Anhui. The agglutination characteristics of the human influenza virus CA/07 (H1N1) and the avian influenza virus AH/05 (H5N1) with known receptor types were also assessed and used as controls.

Gong S., Qi F., Li F., Lv Q., Wang G., Wang S., Jiang J., Wang L., Bao L, & Qin C. (2019). Human-Derived A/Guangdong/Th005/2017 (H7N9) Exhibits Extremely High Replication in the Lungs of Ferrets and Is Highly Pathogenic in Chickens. Viruses, 11(6), 494.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!