Journal: Journal of Cellular and Molecular Medicine

Article Title: Overexpression of sortilin is associated with 5‐FU resistance and poor prognosis in colorectal cancer

doi: 10.1111/jcmm.15752

Figure Lengend Snippet: Sortilin protein expression increases upon 5‐FU treatment in tumours from Nude mice xenografted with CRC cell lines. A, Tumour volume of WiDr and SW620 xenografted Nude mice untreated (CTL) or exposed to 5‐FU (5‐FU) was determined every 3 days from the graft and during the treatment protocol, as described in material and methods ( V = [L × W(L + W)]ᴨ/12). T. I. refers to Treatment Initiation. B, Sortilin protein expression analysis by Western blotting on whole lysates obtained from tumours of untreated xenografted Nude mice and 5‐FU‐treated ones ( n = 5 per conditions). Actin was used as loading control. Histograms represent means of 5 different samples (significant P ‐values are indicated in the graphs * P < .05, ** P < .01, *** P < .001). C, Representative illustration of sortilin protein expression observed by immunohistochemistry (magnification x100) on paraffin‐embedded xenografted Nude mice tumours ( n = 5 per conditions). Histograms represent the immunochemistry intensity score, as described in material and methods

Article Snippet: For immunohistochemistry analysis, primary antibody used was mouse anti‐sortilin (ANT‐016, Alomone Labs).

Techniques: Expressing, Western Blot, Immunohistochemistry

Journal: Journal of Cellular and Molecular Medicine

Article Title: Overexpression of sortilin is associated with 5‐FU resistance and poor prognosis in colorectal cancer

doi: 10.1111/jcmm.15752

Figure Lengend Snippet: High SORT1 expression is associated with poorer survival, DFS and RFS, and sortilin expression is associated with higher CRC tumour grades. SORT1 expression levels by risk group according to survival ( n = 77; A), DFS ( n = 545; B) and RFS ( n = 37; C) of patients suffering from CRC, extracted from ‘SurvExpress’ database. Significant P‐values are indicated in the graphs * P < .05, ** P < .01, *** P < .001. D, Sortilin protein expression, observed by immunohistochemistry (magnification, 50×) on tissue microarrays including seven samples of benign tissue and 126 samples from CRC tumours of different grades ( n = 8, 98, and 20 for grades I, II and III, respectively). Quantification was performed by calculating the immunochemistry intensity score described in materials and methods. E, Sortilin expression in whole‐cell lysates of WiDr and SW620. Actin was used as a loading control

Article Snippet: For immunohistochemistry analysis, primary antibody used was mouse anti‐sortilin (ANT‐016, Alomone Labs).

Techniques: Expressing, Immunohistochemistry

Journal:

Article Title: Improved Hepatic Gene Transfer by Using an Adeno-Associated Virus Serotype 5 Vector

doi: 10.1128/JVI.76.20.10497-10502.2002

Figure Lengend Snippet: Estimation of vector gene copy number in AAV-transduced livers by quantitative PCR (3 months after vector administration). A 350-bp fragment of the hF.IX cDNA as present in the AAV-EF1α-hF.IX vector was coamplified with a 1.1-kb fragment from the endogenous murine HPRT gene using biotinylated primers (20 cycles of 95°C for 1 min, 56°C for 1 min, and 72°C for 2 min, separated on a 2% agarose gel, transferred to a nylon membrane, and visualized with the Southern light detection system from Applied Biosystems. Template for PCR was as follows: 100 ng of genomic DNA extracted from several random pieces of liver and subsequently combined for each individual animal. Each sample column represents an individual animal. standards, linearized plasmid pAAV-EF1α-hF.IX (0.01, 0.1, or 1 pg) mixed with 100 ng of genomic mouse DNA (extracted from untransduced animal); NC, negative control (template, genomic DNA from untransduced animal). Bands were analyzed by densitometric scanning, and intensities were quantitated with NIH Image 6.16 software. Shown is one representative blot. Ratios of band intensities (hF.IX band/mAAT band) are for the blot shown. Gene copy number estimates are average for two experiments.

Article Snippet: A 350-bp fragment of the hF.IX cDNA as present in the AAV-EF1α-hF.IX vector was coamplified with a 1.1-kb fragment from the endogenous murine HPRT gene using biotinylated primers (20 cycles of 95°C for 1 min, 56°C for 1 min, and 72°C for 2 min, separated on a 2% agarose gel, transferred to a nylon membrane, and visualized with the Southern light detection system from Applied Biosystems.

Techniques: Plasmid Preparation, Real-time Polymerase Chain Reaction, Agarose Gel Electrophoresis, Negative Control, Software