Itaq universal sybr green supermix

Deidentified HPLFs, which were isolated from patients with IPF and from non‐disease control (NDC) patients, were provided through the University of Nebraska Medical Center lung tissue biobank with prior approval from the UNMC IRB. All HPLFs were cultured in DMEM with 10% FBS, 1% amphotericin B, 1% l‐glutamine, penicillin (100 U mL⁻¹), and streptomycin (100 µg mL⁻¹) at 37 °C in a humidified chamber with 5% CO₂. MPLFs were isolated from the lungs of mice with BLM‐induced pulmonary fibrosis as described before^[12b] and cultured at 37 °C with 5% CO₂ in high‐glucose DMEM with 10% FBS and Pen‐Strep (100 U mL⁻¹, 100 µg mL⁻¹).
CXCR4, STAT3, col1a1, and col1a2 mRNA levels in the HPLFs and MPLFs were measured by RT‐PCR. Cells were homogenized with TRIzol reagent to isolate total RNA following the protocol. Then the total RNA was converted into cDNA via a High‐Capacity cDNA Transcription kit. The PCR reaction was run on the Rotor‐Gene Q (QIAGEN) using iTaq Universal SYBR Green Supermix.
Deidentified human lung parenchyma slides from NDC and IPF patients were provided by the University of Nebraska Medical Center Lung Transplant Biobank, and the slides were sectioned and analyzed with H&E and Masson's trichrome staining. Immunofluorescence dual‐staining for STAT3 and p‐STAT3, collagen‐1, and α‐SMA (smooth muscle actin) was performed, and slides observed using a confocal microscope.