artificial ace2 substrate mca-apk-dnp  (Bachem)

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Participant demographics and health characteristics.

Article Snippet: 10 μL of neutralized samples was transferred to 96‐well black plates containing 15 μM of fluorogenic ACE2 substrate Mca‐APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS‐60757), to a final concentration of 11.25 μM.

Techniques: Infection, Biomarker Discovery

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Overview of ACE2 serology results and establishment of seropositivity thresholds. (A) Overall view of anti‐ACE2 IgG, IgA, and IgM antibodies of the full cohort, including longitudinal sampling. (B) Quantitative Venn diagram representation of the distribution between IgM, IgA, and IgG ACE2 autoantibodies in the overall cohort of ACE2‐positive individuals. (C) IgG, IgA, and IgM anti‐ACE2 antibodies and associated threshold. IgG and IgA threshold was set at two cycles of 2 standard deviations (SD) from the mean of the presumed negative distribution. IgM threshold was set by a first cycle at 2SD and a second at 1SD from the mean of the presumed negative distribution.

Article Snippet: 10 μL of neutralized samples was transferred to 96‐well black plates containing 15 μM of fluorogenic ACE2 substrate Mca‐APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS‐60757), to a final concentration of 11.25 μM.

Techniques: Sampling

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Influence of clinical features and demographic characteristics on ACE2 autoantibodies prevalence. Odd ratios (OR) and 95% confidence interval (CI) were calculated for demographic characteristics and clinical features for individuals in relation to (A) IgG, (B) IgA and (C) IgM ACE2 autoantibodies. A table with a complete list of OR and 95% CI is available in the Table . The OR (CI95) are shown here on a log10 formatted axis.

Article Snippet: 10 μL of neutralized samples was transferred to 96‐well black plates containing 15 μM of fluorogenic ACE2 substrate Mca‐APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS‐60757), to a final concentration of 11.25 μM.

Techniques:

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Persistence of autoantibodies in serum over time and association with previous SARS‐CoV‐2 infection. (A) Some individuals had one or two follow‐up samples. ACE2 IgG, IgA, IgM is shown longitudinally (Follow up 1 ( n = 120), median time between draws (SD) = 35 days (10), Follow up 2 ( n = 35), median time between draws (SD) = 37 days (6)). (B) IgG, IgA, and IgM ACE2 autoantibodies levels are shown for individuals who never had SARS‐CoV‐2 and for individuals who had serological markers of SARS‐CoV‐2 infection and/or self‐reported a SARS‐CoV‐2 infection. Unpaired two‐sided Wilcoxon test was used to establish statistical significance ( n = 464), p ≤ 0.05 (*), p ≤ 0.01 (**), p ≤ 0.001 (***), p ≤ 0.0001 (****).

Article Snippet: 10 μL of neutralized samples was transferred to 96‐well black plates containing 15 μM of fluorogenic ACE2 substrate Mca‐APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS‐60757), to a final concentration of 11.25 μM.

Techniques: Infection

Journal: The FASEB Journal

Article Title: Autoantibodies targeting angiotensin‐converting enzyme 2 are prevalent and not induced by SARS ‐ CoV ‐2 infection

doi: 10.1096/fj.202402694R

Figure Lengend Snippet: Effect of ACE2 autoantibodies on ACE2 enzymatic activity and SARS‐CoV‐2 spike‐ACE2 binding. (A) ACE2 autoantibodies levels (IgG, IgA, IgM) in a subset of samples for which ACE2 enzymatic activity was assessed. (B) Levels of ACE2 enzymatic activity in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies in contrast to the individuals who had no detectable ACE2 autoantibodies. (C) Levels of ACE2 enzymatic activity in individuals who self‐reported a SARS‐CoV‐2 infection and/or had serological markers of previous infection. (D) ACE2 autoantibodies levels in a subset of samples in which ACE2 and spike interaction was assessed. (E) Anti‐Spike IgG antibodies levels pre‐ and post‐depletion in sera. (F) % inhibition representing the ability of the sera to inhibit spike and ACE2 interaction pre‐ and post‐depletion. (G) % inhibition representing the ability of the sera to inhibit spike and ACE2 interaction in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies above threshold in contrast to the individuals who had no detectable ACE2 autoantibodies. (H) Linear correlation between the ability of sera to inhibit Spike and ACE2 binding and the levels of Spike IgG antibodies in depleted sera. Unpaired two‐sided Wilcoxon test was used to establish statistical significance where applicable, p ≤ 0.05 (*), p ≤ 0.01 (**), p ≤ 0.001 (***), p ≤ 0.0001 (****).

Article Snippet: 10 μL of neutralized samples was transferred to 96‐well black plates containing 15 μM of fluorogenic ACE2 substrate Mca‐APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS‐60757), to a final concentration of 11.25 μM.

Techniques: Activity Assay, Binding Assay, Infection, Inhibition

Journal: medRxiv

Article Title: Autoantibodies Targeting Angiotensin Converting Enzyme 2 Are Prevalent and Not Induced by SARS-CoV-2 Infection

doi: 10.1101/2024.11.18.24317488

Figure Lengend Snippet: A) overall view of anti-ACE2 IgG, IgA, and IgM antibodies of the full cohort including longitudinal sampling. B) Quantitative Venn diagram representation of the distribution between IgM, IgA, and IgG ACE2 autoantibodies in the overall cohort of ACE2-positive individuals. C) IgG, IgA, and IgM anti-ACE2 antibodies and associated threshold. IgG and IgA threshold was set at two cycles of 2 standard deviations (SD) from the mean of the presumed negative distribution. IgM threshold was set by a first cycle at 2SD and a second at 1SD from the mean of the presumed negative distribution.

Article Snippet: 10uL of neutralized samples were transferred to 96-well black plates containing 15 uM of fluorogenic ACE2 substrate Mca-APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS-60757), to a final concentration of 11.25 uM ( ).

Techniques: Sampling

Journal: medRxiv

Article Title: Autoantibodies Targeting Angiotensin Converting Enzyme 2 Are Prevalent and Not Induced by SARS-CoV-2 Infection

doi: 10.1101/2024.11.18.24317488

Figure Lengend Snippet: Odd ratios (OR) and 95% confidence interval (CI) were calculated for demographi characteristics and clinical features for individuals in relation to A) IgG, B) IgA and C) IgM ACE2 autoantibodies. A table with a complete list of OR and 95% CI is available in the supplementary material (Supp. ). The OR (CI95) are shown here on a log10 formatted axis.

Article Snippet: 10uL of neutralized samples were transferred to 96-well black plates containing 15 uM of fluorogenic ACE2 substrate Mca-APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS-60757), to a final concentration of 11.25 uM ( ).

Techniques:

Journal: medRxiv

Article Title: Autoantibodies Targeting Angiotensin Converting Enzyme 2 Are Prevalent and Not Induced by SARS-CoV-2 Infection

doi: 10.1101/2024.11.18.24317488

Figure Lengend Snippet: A) Some individuals had one or two follow-up samples. ACE2 IgG, IgA, IgM is shown longitudinally (Follow up 1 (n=120), median time between draws (SD) = 35 days , Follow up 2 (n=35), median time between draws (SD) = 37 days ). B) IgG, IgA, IgM ACE2 autoantibodies levels are shown for individuals who never had SARS-CoV-2 and for individuals who had serological markers of SARS-CoV-2 infection and/or self-reported a SARS-CoV-2 infection. Unpaired two-sided Wilcoxon test was used to establish statistical significance (n=464), * p< 0.05, ** p< 0.01, ***p<0.001, ****p<0.0001.

Article Snippet: 10uL of neutralized samples were transferred to 96-well black plates containing 15 uM of fluorogenic ACE2 substrate Mca-APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS-60757), to a final concentration of 11.25 uM ( ).

Techniques: Infection

Journal: medRxiv

Article Title: Autoantibodies Targeting Angiotensin Converting Enzyme 2 Are Prevalent and Not Induced by SARS-CoV-2 Infection

doi: 10.1101/2024.11.18.24317488

Figure Lengend Snippet: A) ACE2 autoantibodies levels (IgG, IgA, IgM) in a subset of samples for which ACE2 enzymatic activity was assessed. B) Levels of ACE2 enzymatic activity in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies in contrast to the individuals who had no detectable ACE2 autoantibodies. C) Levels of ACE2 enzymatic activity in individuals who self-reported a SARS-CoV-2 infection and/or had serological markers of previous infection. D) ACE2 autoantibodies levels in a subset of samples in which ACE2 and spike interaction was assessed. E) Anti-Spike IgG antibodies levels pre-and post-depletion in sera. F) % inhibition representing the ability of the sera to inhibit Spike and ACE2 interaction pre-and post-depletion. G) % inhibition representing the ability of the sera to inhibit Spike and ACE2 interaction in individuals who had IgG and/or IgA and/or IgM ACE2 autoantibodies above threshold in contrast to the individuals who had no detectable ACE2 autoantibodies. H) Linear correlation between the ability of sera to inhibit Spike and ACE2 binding and the levels of Spike IgG antibodies in depleted sera. Unpaired two-sided Wilcoxon test was used to establish statistical significance where applicable, * p< 0.05, ** p< 0.01, ***p<0.001, ****p<0.0001.

Article Snippet: 10uL of neutralized samples were transferred to 96-well black plates containing 15 uM of fluorogenic ACE2 substrate Mca-APK(Dnp) (Anaspec, Fremont, CA, USA, cat. AS-60757), to a final concentration of 11.25 uM ( ).

Techniques: Activity Assay, Infection, Inhibition, Binding Assay

Journal: Journal of Molecular Medicine (Berlin, Germany)

Article Title: Novel high-yield potato protease inhibitor panels block a wide array of proteases involved in viral infection and crucial tissue damage

doi: 10.1007/s00109-024-02423-x

Figure Lengend Snippet: Potato PI fractions inhibit a range of proteases relevant to virus infection and tissue damage

Article Snippet: Concentrations between 100 nM and 1 μM were used for fractions 1–5 and incubated for 6 h at 37 °C, whereafter 2 µL of the ACE2 protease substrate solution (Abcam, UK #111,750) was added.

Techniques: Virus, Infection

Journal: Journal of Molecular Medicine (Berlin, Germany)

Article Title: Novel high-yield potato protease inhibitor panels block a wide array of proteases involved in viral infection and crucial tissue damage

doi: 10.1007/s00109-024-02423-x

Figure Lengend Snippet: Inhibition of ACE2 activity in Vero E6 cells by the PI fractions. A – E Proteolytic activity of ACE2 measured from fractions 1–5 in Vero E6 cells. All individual experiments were performed in triplicate. Significance was tested using Students’ test. p values are indicated as **** p < 0.0001, *** p < 0.001, ** p < 0.01, and * p < 0.05

Article Snippet: Concentrations between 100 nM and 1 μM were used for fractions 1–5 and incubated for 6 h at 37 °C, whereafter 2 µL of the ACE2 protease substrate solution (Abcam, UK #111,750) was added.

Techniques: Inhibition, Activity Assay

Journal: Journal of Molecular Medicine (Berlin, Germany)

Article Title: Novel high-yield potato protease inhibitor panels block a wide array of proteases involved in viral infection and crucial tissue damage

doi: 10.1007/s00109-024-02423-x

Figure Lengend Snippet: Inhibitory activity of potato PI fractions against enzymes in viral replication and tissue damage

Article Snippet: Concentrations between 100 nM and 1 μM were used for fractions 1–5 and incubated for 6 h at 37 °C, whereafter 2 µL of the ACE2 protease substrate solution (Abcam, UK #111,750) was added.

Techniques: Activity Assay