Sc 57351

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Sc-57351 is a laboratory product offered by Santa Cruz Biotechnology. It is a piece of equipment designed for scientific research and analysis. The core function of Sc-57351 is to facilitate experimental procedures in a laboratory setting.

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3 protocols using sc 57351

Comprehensive Antibody Validation Protocol

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Antibodies against Gbp2 (ab203238), GFAP (ab53554, ab7260), LCN2 (ab63929), LRP1 (ab92544), MAG (ab89780), MBP (ab40390), NF200 (ab7795), PLP (ab28486) and S100A10 (ab76472) were purchased from Abcam, UK; antibody against CD31 (550274) was purchased from BD Biosciences, USA; antibodies against GFAP (3670S), GFP (2955S, 2956S), p38 (9212S), pp38 (4511S) and β-actin (8457S) were purchased from Cell Signaling Technology, USA; antibodies against CC1 (OP80), dMBP (AB5864) and Olig2 (AB9610) were purchased from Millipore, USA; antibodies against C3d (AF2655), Galectin3 (AF1197), LAMP1 (AF4320) and LCN2 (AF1857) were purchased from R&D Systems, USA; antibodies against Gbp2 (sc-166960) and LRP1 (sc-57351) were purchased from Santa Cruz Biotechnology, USA; antibody against pLRP1 (PA5-101013) was purchased from Thermo Fisher, USA; antibody against Iba1 (019-19741) was purchased from Wako, Japan. All antibodies were used at a dilution of 1:50 to 1:2000 for immunofluorescence and 1:800 to 1:5000 for immunoblotting according to the manufacturer’s instructions. Alexa Fluor 488, 594 and 647 conjugated secondary antibodies were purchased from Jackson, USA.

Wan T., Zhu W., Zhao Y., Zhang X., Ye R., Zuo M., Xu P., Huang Z., Zhang C., Xie Y, & Liu X. (2022). Astrocytic phagocytosis contributes to demyelination after focal cortical ischemia in mice. Nature Communications, 13, 1134.

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LRP1 Expression Analysis in MVNs

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Expression analysis of LRP1 was performed using a Proteinsimple automatic western assay as previously described (73) (74) (74)after fixation with PFA. Devices was separated from the glass coverslip using a razor blade, and different regions of the MVNs in fibrin gel were collected: 4 ROIs in the control MVNs without GBM spheroids, and in the MVNs with GBM spheroids, 2 ROIs near and 2 ROIs far from the spheroid.; n=5-6 devices were employed for each condition. Samples were incubated in lysis buffer comprising 10mL of 1X buffer (9803S, Cell Signaling Technologies), 1 µL Benzonase Nuclease (E8263, Millipore Sigma), one tablet of protease inhibitor cocktail (11836170001, Millipore Sigma), and stored at -80 °C. LRP1 signal (sc-57351, Santa Cruz Biotechnology) was normalized to CD31 (ab32457, Abcam) or β-actin (926-42210, Li-Cor) using Compass software v5.0. The output of this automatic western assay is not a standard blot but rather a chemiluminescence spectrum; representative uncropped raw data for LRP1 signal in one device is included in Figure S1.

Straehla J.P., Hajal C., Safford H.C., Offeddu G.S., Boehnke N., Dacoba T.G., Wyckoff J., Kamm R.D., & Hammond P.T. (2021). A predictive microfluidic model of human glioblastoma to assess trafficking of blood-brain barrier penetrant nanoparticles.

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Multimodal Immunofluorescence Imaging of Cellular Markers

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Devices were fixed, permeabilized, blocked prior to staining. Protein visualization was achieved by staining the fixed devices with anti-CD31 (ab3245, Abcam), anti-LRP1 (sc-57351, Santa Cruz Biotechnology), and anti-ZO-1 (61-7300, ThermoFisher Scientific) at 1:200 in PBS, overnight at 4C on a shaker. Secondary antibodies were used at 1:200 in PBS (568 goat anti-rabbit A-11011, or 633 goat anti-mouse A-21052, Invitrogen) and DAPI (D1306, Invitrogen) at 1:1000. Additional staining details are in the supplemental methods. Images were acquired with a confocal laser scanning microscope (FV-1200, Olympus).
For in vivo samples, brains were formalin fixed and paraffin embedded before staining with cleaved caspase-3 (CC3 Rabbit Mab, 1:800 #9664L [D175] Cell Signaling Technology) and rabbit polymer secondary (Biocare Medical # RMR 622L). Quantification of CC3 staining was performed in QuPath v0.2.3 (Queen's University, Belfast, Northern Ireland) using QuPath's build-in 'Positive cell detection' (72) with three regions of interest of the same size manually placed per tumor section.

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