Human Pro Collagen I alpha 1 Antibody

Detection of Human N-Pro Collagen I by Immunoprecipitation.

Human N-Pro Collagen I was immunoprecipitated from 500 μg of IMR‑90 human lung fibroblast cell line lysates with 5 μg of Sheep Anti-Human N-Pro Collagen I Antigen Affinity-purified Polyclonal Antibody (Catalog # AF6220). The N-Pro Collagen I-antibody complexes were absorbed using Protein A or Protein G. Immunoprecipitated human N-Pro Collagen I was detected by Western blot using 1 μg/mL of Mouse Anti-Human N-Pro Collagen I Monoclonal Antibody (MAB6220) under reducing conditions and using Western Blot Buffer Group 1.

Detection of Human Collagen I  alpha 1 by Simple Western^TM.

Simple Western lane view shows lysates of IMR‑90 human lung fibroblast cell line, loaded at 0.2 mg/mL. A specific band was detected for Collagen I  alpha 1 at approximately 232 kDa (as indicated) using 10 µg/mL of Sheep Anti-Human Pro Collagen I  alpha 1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF6220) followed by 1:50 dilution of HRP-conjugated Anti-Sheep IgG Secondary Antibody (HAF016). This experiment was conducted under reducing conditions and using the 66-440 kDa separation system.

Detection of Pro-Collagen I alpha 1 by Immunohistochemistry

Comparison of gene expression in a scRNA-seq and Spatial Transcriptomics analysis of hESC-derived DA neuronsExpression of genes associated with dopaminergic neurons (SLC6A3, TH), astrocytes (AQP4, SOX9), and VLMCs (COL1A1) in (A) scRNA-seq of previously analyzed transplant, (B) ST analysis of grafted tissue (transplant area is delineated in the medial striatum), and (C) ST analysis of intact substantia nigra. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37251982), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Pro-Collagen I alpha 1 by Immunocytochemistry/ Immunofluorescence

Decreased ECM collagens and fibronectin in KC keratocyte cultures. Representative images of Sirius red and fast green stained cultures (20x) in (A) 2 separate DN and (B) 2 separate KC cultures, scale bar: 50 μm. (C) Cell associated hydroxyproline quantification for total collagenous protein, (D) media associated hydroxyproline. Each point representing means of three measurements from individual cultures, n = 6. Statistical significance was assessed using the Student's t-test; *P < 0.05. (E) Cell immunoblot of COL1A1 and GAPDH from 14-day cultures. (F) Corresponding Immunoblot of COL1A1 released into the media over 3 days (days 11–14) with total protein for normalization. Quantitative PCR of (G) COL1A1 and (H) COL5A1 transcription relative to POLR2A. Points represent mean of individual donor cultures and shown as mean ± SEM; statistical significance was assessed using the Student's t-test; *P < 0.05, **P < 0.01. (I, J) Immunohistochemical localization of COL1A1, COL5A1 and FN1 in DN cultures and KC cultures. Green: protein of interest, Red: F-actin, Blue: DAPI. Scale bar: 50 μm. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30029277), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Pro-Collagen I alpha 1 by Western Blot

Increased MMP2 and decrease in collagen maturation related proteins in KC keratocytes. (A) Gelatin zymography of day 14 media (days 11–14) with bands corresponding to 65 kDa latent MMP2, and 60 kDa active MMP2 in 6 DN and 6 KC patients. (B) Western blots of HSP47, CTAGE5 and CALR and GAPDH as internal control. (C–E) Quantification of HSP47, CTAGE5, CALR, points represent mean of individual donor cultures and range is standard error of mean. Values are shown as mean ± SEM; statistical significance was assessed using the Student's t-test; *P < 0.05, **P < 0.01, ***P < 0.005. (F–H) Quantitative PCR of BMP1, PCOLCE, ADAMTS2, using 2-delta delta cT relative to POLR2A, n ≥ 9 biological repeats. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/30029277), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Pro-Collagen I alpha 1 by Western Blot

Overexpressing BMP1 in PANC1 cells suppresses tumor growth and metastasis.a–e Orthotopic injection of PANC1 cells (a) overexpressing BMP1 with lenti-BMP1 (b) resulted in reduced primary tumor weight (c), and reduced liver metastasis load after normalization to primary tumor weight (d). Representative lung and liver metastasis images are shown in e with both zsgreen signal and human LMNA IHC to highlight cancer cells. Note that PANC1 cells are not metastatic to the lungs. Mouse numbers are 8 and 6 (left to right). f, g Cultured lenti-BMP1 PANC1 cells deposit 4x more alpha 1(I) compared to control cells (f) although COL1A1 RNA level is comparable (g). f Shows images representative of three independent experiments with beta -actin as a sample processing control. The quantified WB signal is represented as mean ± SD. h, i IHC on xenograft tumor sections showed increased intensities in both cancer-cell- and stromal-derived ColI staining upon BMP1 overexpression in the PANC1 cells (h). The intensities were quantified in i (N = 7 and 6, left to right). j, k WB on enriched ECM samples from xenograft PANC1 tumors showed increased cancer-cell-derived (human) and stromal-derived (mouse) ColI deposition in lenti-BMP1 tumors. See Methods section for antibodies used. k is the quantification of the band intensities of four tumors (two were shown in j and two came from an independent repeat). l The normalized intensity of protein Hypro to either protein proline or aspartic acid in xenograft tumors suggested an increase in overall collagen upon BMP1 overexpression in PANC1 cells (N = 3). All p-values come from two-tailed Student’s t tests. All columns are represented by mean ± SD. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/33879793), licensed under a CC-BY license. Not internally tested by R&D Systems.