Recombinant Mouse Ace2 protein, His-tagged

• Cat.No.: Ace2-761M
• Product Overview: Recombinant Mouse Ace2 protein(Gln 18 - Thr 740), fused with His tag, was expressed in HEK293.

Specification

• Species: Mouse
• Source: HEK293
• Tag: His
• Protein Length: Gln 18 - Thr 740
• Form: Supplied as 0.2 μm filtered solution in 50 mM Tris, 150 mM NaCl, Arginine, pH7.5 with glycerol as protectant.
• Molecular Mass: This protein carries a polyhistidine tag at the N-terminusThe protein has a calculated MW of 85.4 kDa. The protein migrates as 90-110 kDa under reducing (R) condition (SDS-PAGE) due to glycosylation.
• Endotoxin: Less than 1.0 EU per μg by the LAL method.
• Purity: >95% as determined by SDS-PAGE. >90% as determined by SEC-MALS.
• Storage: For long term storage, the product should be stored at lyophilized state at -20°C or lower.Please avoid repeated freeze-thaw cycles. This product is stable after storage at: -20°C to -70°C for 12 months in lyophilized state; -70°C for 3 months under sterile conditions after reconstitution.
• Reconstitution: It is recommended that sterile water be added to the vial to prepare a stock solution of 0.2 ug/ul. Centrifuge the vial at 4°C before opening to recover the entire contents.

Gene Information

• Gene Name: Ace2 angiotensin I converting enzyme (peptidyl-dipeptidase A) 2 [ Mus musculus ]
• Official Symbol: Ace2
• Synonyms: ACE2; angiotensin I converting enzyme (peptidyl-dipeptidase A) 2; angiotensin-converting enzyme 2; ACE-related carboxypeptidase; angiotensin I converting enzyme 2; 2010305L05Rik
• Gene ID: 70008
• mRNA Refseq: NM_001130513
• Protein Refseq: NP_001123985

Citation

CXCL12 and CXCL13 Cytokine Serum Levels Are Associated with the Magnitude and the Quality of SARS-CoV-2 Humoral Responses

Journal: Viruses    PubMed ID: 36560669    Data: 2022/11/28

Authors: Alessandra Noto, Victor Joo, Caijun Sun

Article Snippet:Control wells were included on each 96-well plate that included beads alone, matching the serum dilutions of a control pool of pre-COVID-19 pandemic healthy human sera (BioWest human serum AB males; VWR) and a positive control of commercial anti-Spike blocking antibody (SAD-S35, from ACRO Biosystem, Newark, DE, USA ) or recombinant-produced REGN10933 neutralizing antibody, discovered and marketed by Regeneron and tested in a concentration response.pool of pre-COVID-19 pandemic healthy human sera (BioWest human serum AB males; VWR) and a positive control of commercial anti-Spike blocking antibody (SAD-S35, from ACRO Biosystem, Newark, DE, USA ) or recombinant-produced REGN10933 neutralizing antibody, discovered and marketed by Regeneron and tested in a concentration response. ... The plates were agitated on a plate shaker for 60 min, then the ACE-2 mouse Fc fusion protein (either Creative BioMart or produced by the EPFL Protein Production and Structure Core Facility) was then added to each well, at a final concentration of 1 μg/mL, and agitated for a further 60 min.. The beads were then washed with the magnetic plate washer, then anti-mouse IgG-PE secondary antibody (OneLambda, Thermo Fisher) was added at a 1/100 dilution, with 50 μL per well.The beads were then washed with the magnetic plate washer, then anti-mouse IgG-PE secondary antibody (OneLambda, Thermo Fisher) was added at a 1/100 dilution, with 50 μL per well.

Antibody responses against the Spike protein in ICU, non-ICU, and non-hospitalized individuals. Luminex beads, coupled with Spike proteins, were used to monitor the IgG binding antibody response; neutralization was measured by a surrogate Spike/ACE-2 inhibition binding assay in the sera, from ICU (N = 31), non-ICU (N = 41), and non-hospitalized (N = 23) individuals at 6 months post-SARS-CoV-2 infection. The proportion of individuals with detectable ( A ) IgG ( C ) neutralizing antibodies in the three study groups. ( B ) Levels of IgG-binding antibody responses: MFI signals for serum antibody-binding were expressed as ratios, compared to a negative-control pool of pre-COVID-19 pandemic human serum, tested in parallel. ( D ) IC50 dilutions of the Spike/ACE2 surrogate neutralization assay. p -values were obtained using chi-square and a one-way ANOVA (Kruskal–Wallis test), followed by a Mann–Whitney test. Red stars indicate statistical significance (** = p < 0.001; *** = p < 0.001); ns = not significant.

A multiplexed high-throughput neutralization assay reveals a lack of activity against multiple variants after SARS-CoV-2 infection

Journal: medRxiv    Data: 2021/4/13

Authors: Fenwick Craig, Turelli Priscilla, Trono Didier

Article Snippet:PrePrint: Control wells were included on each 96-well plate that included beads alone, matching serum dilutions of a control pool of pre-COVID-19 pandemic healthy human sera (BioWest human serum AB males; VWR) and a positive control commercial anti-Spike blocking antibody (SAD-S35 from ACRO Biosciences) or recombinant produced REGN10933 neutralizing antibody discovered and marketed by Regeneron tested in a concentration response.serum dilutions of a control pool of pre-COVID-19 pandemic healthy human sera (BioWest human serum AB males; VWR) and a positive control commercial anti-Spike blocking antibody (SAD-S35 from ACRO Biosciences) or recombinant produced REGN10933 neutralizing antibody discovered and marketed by Regeneron tested in a concentration response. ... Plates were agitated on a plate shaker for 60 minutes then the ACE2 mouse Fc fusion protein (Creative BioMart or produced by EPFL Protein Production and Structure Core Facility) was then added to each well at a final concentration of 1 μg/ml and agitated for a further 60 minutes.. Beads were then washed on the magnetic plate washer and anti-mouse IgG-PE secondary antibody (OneLambda ThermoFisher) was added at a 1/100 dilution with 50μl per well.Beads were then washed on the magnetic plate washer and anti-mouse IgG-PE secondary antibody (OneLambda ThermoFisher) was added at a 1/100 dilution with 50μl per well.

( a ) Schematic outline of the S 3 /ACE2 neutralization assay. Anti-SARS-CoV-2 serum antibodies are monitored for their capacity in blocking the S 3 /ACE2 interaction. ACE2 binding to S protein was detected though use of a fluorescently labeled secondary antibody and the signal intensities are inversely proportional to the neutralizing potential of the anti-S protein antibodies. ( b ) Serum dilution IC 50 values were calculated for 104 healthy adult donor samples collected prior to November 2019 (pre-COVID-19 pandemic). The mean IC50 values and SD were used to establish a lower limit cutoff of 50 indicated by the dashed line (mean IC 50 of 12.5 + 4 × 9.0 SD = 50 serum dilution). ( c ) Representative concentration response curves for ten healthy donor serum samples and ( d ) ten SARS-CoV-2 seropositive donors with varying levels of anti-S protein IgG antibody. Mean ± s.d. shown in graph b . S protein / ACE2 structure was generated with PDB 7a98.

( a ) Cross-validation studies between S 3 /ACE2 surrogate neutralization assay and the “gold standard” SARS-CoV-2 cytopathic effect (CPE) neutralization assay. Seropositive donors (n=206) with varying levels of anti-S protein antibodies were selected for comparison of the assays. Donors consisted of COVID-19 hospitalized patients (n=31), symptomatic infected donors with RT-PCR-documentation (n=64) and other seropositive donors identified through random sampling, volunteers or contact with a confirmed SARS-CoV-2 infected individual in a Swiss population serological survey (n=111). ( b ) Correlation between the live SARS-CoV-2 virus cytopathic effect and S protein pseudotyped neutralization assays. A group of 74 samples from S protein seropositive donors were compared in the live virus CPE and S protein pseudotyped virus cell based neutralization assays. Correlation between the two neutralization assays is represented by the black dashed line and the 20 serum dilution IC 50 cutoff for positivity in the CPE assay is shown with the blue dashed line.