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Fig. 1 | Respiratory Research

Fig. 1

From: ColdZyme® protects airway epithelia from infection with BA.4/5

Fig. 1

ColdZyme® mouth spray protects primary human airway epithelial (HAE) cells from SARS-CoV-2 BA.1 and BA.4/5 infection and innate immune activation. Visualization of virus binding (SARS-CoV-2 S1/N, orange) and complement (C3-FITC, green) in SARS-CoV-2 infected 3D pseudostratified epithelia. Pseudostratified epithelia were apically treated with ColdZyme® mouth spray prior exposure to SARS-CoV-2 (MOI 0.01). On 3 dpi, filters were fixed, stained for höchst (blue), SARS-CoV-2 S1/N (orange), complement C3 (green) and WGA (red) and then analysed by HCS. a Overview on one half of a Transwell filter of solvent/ColdZyme®-controls (panel 1), BA0.1- or BA0.4/5-infected (panel 2 and 4), ColdZyme-pre-treated and BA0.1 or BA0.4/5-infected (panel 3 and 5) HAE cultures using the 5X magnification and showing all markers or b complement C3 (green) and SARS-CoV-2 (N/S) (orange) markers. One representative filter half is illustrated. Scale bars represent 2 mm. c Z-stacks of uninfected (UI, panel 1), BA.1- or BA.4/5-infected (panel 2 and 4), ColdZyme-pre-treated and BA.1 or BA.4/5-infected (panel 3 and 5) HAE cultures were analyzed using the Operetta CLS HCS and the 63XWATER objective. Cells were stained using C3-FITC (green) as indicator for innate immune activation, SARS-CoV-2-S1/N-Alexa594 (orange) for virus detection, höchst for imaging nuclei (blue) and WGA for staining lectins (red). High IC C3 mobilization was monitored in BA.1- and BA.4/5-infected cultures, while no virus and low C3 signals were detected in UI (panel 1) and ColdZyme®/BA.1 or BA.4/5- (panel 3 and 5) cultures. Scale bars represent 50 μm and three independent experiments were performed. d Tissue destruction upon viral infection (BA0.1, panel 1; BA0.4/5, panel 3) and rescue by single ColdZyme® application prior infection (BA.1, panel 2; BA.4/5, panel 4) are shown. Scale bars represent 50 μm and three independent experiments were performed. e More than 1000 cells per condition (UI, BA.1, BA.1 + ColdZyme), BA.4/5, BA.4/5 + ColdZyme) were analyzed for their expression of C3, where up to 38% for BA.1 and 60% for BA.4/5 of the analyzed cells were stained positive for C3. Only 10–18% of C3 signal were detected in UI or ColdZyme/BA.1 or BA.4/5-exposed cells. Statistical significances were analyzed with GraphPad Prism software using One-way ANOVA and Tukey´s post test

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