Chemiluminescence method

Cells lysed in ice‐cold RIPA buffer (Beyotime, China) supplemented with 10 nM PMSF. The protein samples of equal amount were resolved on 10% SDS polyacrylamide gels and transferred to polyvinylidene fluoride membranes at 100 V for 2.5 hours. 5% fat‐free milk in TBST was used to block the membrane, followed by adding primary antibodies (Abcam, Cambridge) (anti‐LDHA, 1: 500) and incubation at 4°C overnight. Secondary antibodies (1:5000) were added and incubated for 2 hours at room temperature. The protein bands were visualized using the chemiluminescence method (Millipore, MA). Image J software (National Institutes of Health, Bethesda) was used to analyze the protein expression levels. GAPDH (1:1000) was used as the control.

Extracted proteins from tissues and cells using Radio Immunoprecipitation Assay lysis buffer (RIPA, R0010) from Solarbio Life Sciences (Beijing, China). The BCA reagent kit from Thermo Scientific (23225, Waltham, USA) was used to detect protein concentration. Protein lysates were loaded on a 10% tris-glycine gel and transferred to a 0.22 μm polyvinylidene difluoride membrane. The primary antibodies (Supplementary Table 3) used to detect protein bands in the membranes included HIF-1α, YAP1, phosphorylated (p)-YAP(S127), Collagen type I alpha 1 (Col1A), mitochondrial oxide phosphorylation complex cocktails (OXPHOS), HIF-2α, α-smooth muscle actin (α-SMA), tubulin from Cell Signaling Technology (Danvers, USA) and β-actin from Sigma-Aldrich (St. Louis, US). The anti-rabbit immunoglobin G HRP-linked antibody and anti-mouse immunoglobin G horseradish from Cell Signaling Technology (Danvers, USA). The chemiluminescence method from Millipore Corporation was used to display the protein bands of western blot. The protein bands were visualised using an Amersham Imager 600 (GE, USA) and were quantified using densitometry analysis (Image J x64 software, NIH).

Samples containing equal amounts of protein were denatured in SDS sample buffer, subjected to SDS-PAGE, transferred onto a PVDF membrane, and applied to immunoblot analysis. Protein bands on immunoblots were visualized by a chemiluminescence method (Millipore) and an imaging documentation system (ImageQuant LAS 4000, GE healthcare). Images were analyzed using ImageJ. Primary antibodies against CCNY (Proteintech group), GFP (Roche), GluA1 (a gift from Michael Ehlers, Pfizer Neuroscience), phospho-GluA1 (S845) (Thermo scientific), PSD-95 (Thermo scientific, 7E3-1B8), Synaptophysin (Synaptic Systems), Prox1 (Proteintech group), Ctip2 (Genetex), Py (a gift from D.T.S. Pak, Georgetown Univ.) or β-tubulin (Abcam) were used.

The C. albicans cell pellet was harvested by centrifugation, and cell walls were prepared as described elsewhere44 (link). Briefly, the cell pellet was washed 3 times in PBS (pH 7.4) and then centrifuged, then ground to a fine powder in liquid nitrogen at the presence of a protease inhibitor cocktail. Isolated cell walls were washed 3 times with MilliQ water and then boiled for 10 min in SDS extraction buffer [150 mM NaCl, 2% (w/v) SDS, 100 mM Na-EDTA, 100 mM beta-mercaptoethanol, 50 mM Tris/HCl, pH 7.8]. Subsequently, they were washed again with MilliQ water and stored at −20 °C until required.
Equal amounts of proteins of rSap2, cell walls and total cell lysates were separated by SDS-PAGE (10%), and transferred to a PVDF membrane (Millipore, Germany) using a blotting system (Bio-Rad, USA) for 1 h at 90 V. The membrane was incubated with the scFv-phages (1 × 10¹⁰ pfu), anti-rSap2 pAb and KM13 phages respectively, and subsequently blotted with different secondary antibodies, i.e., anti-phage g3p antibody for phages and goat anti-rabbit IgG for anti-rSap2 pAb. Detection was achieved with the chemiluminescence method according to the manufacturer’s instructions (Millipore, Germany), and visualized by using the ECL detection system (Tanon 5500, Shanghai, China).