Collected cells (GES, AGS, BGC-823, HGC-27, and MKN-28) were lysed on ice with phosphatase and protein inhibitors in RIPA buffer for 30 min, then centrifuged at 13,000 rpm for 15 min. The cell debris was removed, the supernatant was obtained using a pipette, a certain amount of 5× loading buffer was added, and samples were heated at 100 °C for 10 min. The BCA protein analysis kit (Thermo Scientific, Waltham, MA, USA) was used to quantitatively detect the protein concentration. The protein was dissolved using 12% SDS-PAGE, the protein was transferred to a PVDF membrane at low temperatures, 5% skimmed milk powder was used for blocking for 2 h, and the membrane was incubated with the primary antibody overnight at 4 °C (FBLN5: 1:1000, ABclonal, USA; β-Tubulin: 1:1000, ABclonal, USA). The membrane was incubated for 1 h the next day with a horseradish peroxidase-labeled secondary antibody (1:5000). The ECL kit (Thermo Scientific, USA) was used to assess the expression of individual proteins. The experiment was conducted in triplicate (Figure S1 of Supplementary Materials ).
Fbln5
Manufactured by ABclonal
Sourced in United States, China
FBLN5 is a protein that plays a role in the extracellular matrix and has been implicated in various biological processes. It is commonly used in research applications. Detailed information on its specific functions and intended uses is not available in a concise, unbiased, and factual format.
Automatically generated - may contain errors
Lab products found in correlation
Ecl kit, by Thermo Fisher Scientific (1 mentions)
Bca protein analysis kit, by Thermo Fisher Scientific (1 mentions)
β tubulin, by ABclonal (1 mentions)
Loxl1, by Abcam (1 mentions)
Vimentin, by Abcam (1 mentions)
Gapdh, by Abcam (1 mentions)
P akt, by Abcam (1 mentions)
E cadherin, by Abcam (1 mentions)
N cadherin, by Abcam (1 mentions)
Flag tag (flag), by ABclonal (1 mentions)
Anti loxl1, by Abcam (1 mentions)
P erk, by Abcam (1 mentions)
3 protocols using fbln5
1
Protein Expression Analysis of Cell Lines
2
Multicolor Immunofluorescence Imaging of Tumor Tissues
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Double and triple immunofluorescent staining of tumor tissues was carried out, referring to the instructions. Briefly, the slides of tissues were prepared through deparaffinization and a quenching procedure of endogenous peroxidase, and the slides were incubated with primary antibodies against LOXL1 (Abcam), FBLN5 (ABclonal), or the αvβ3 integrin (Abcam), overnight at 4°C. After washed three times with phosphate-buffered saline (PBS) and blocked with goat serum, the slides were incubated with different fluorescein-labeled secondary antibodies. Slides were viewed and photographed with a confocal microscope (Leica).
3
Protein Expression Analysis Protocol
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Protein samples from tissues, cell lysates, or supernatants were lysed and denaturalized for analysis. The following procedures were performed as described previously:27 (link) anti-LOXL1, GAPDH, N-cadherin, E-cadherin, vimentin, AKT, pAKT, Erk, and pErk (Abcam, MA, USA). Flag-tag, FBLN5, FAK pY861, FAK pY397, and FAK (ABclonal, China) antibodies were used as primary antibodies to examine protein expressions. An ECL kit (Pierce; Thermo Fisher Scientific, Rockford, IL, USA) was used to detect specific signals.
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