Total protein was first extracted using SDS-PAGE and then transferred to a PVDF membrane (Thermo, USA). Afterwards, the PVDF membrane was incubated with 1:1000 dilution of antibody: TRIM47 (ab155549, Abcam, USA), BAP1 (ab199396, Abcam, USA), P21 (ab188224, Abcam, USA), P53 (ab131442, Abcam, USA), SRC (ab47405, Abcam, USA), MET (ab51067, Abcam, USA), and c-Myc (ab39688,Abcam, USA). The membrane was washed and incubated with a 1:2000 dilution of horseradish peroxidase-conjugated goat anti-rabbit (Santa Cruz, USA). SuperSignalMT West Puico PLUS (Termo Scientific, USA) was used to develop the blot, using b-actin (ab8226, Abcam, USA) as the loading control. All experiments were performed in triplicate.
Ab155549
Manufactured by Abcam
Sourced in United States
Ab155549 is a lab equipment product. It is a solid support for protein purification.
Automatically generated - may contain errors
Lab products found in correlation
Pvdf membrane, by Thermo Fisher Scientific (2 mentions)
Ab51067, by Abcam (2 mentions)
Ab39688, by Abcam (2 mentions)
Ab199396, by Abcam (2 mentions)
β actin, by Abcam (2 mentions)
Ab131442, by Abcam (2 mentions)
Ab188224, by Abcam (2 mentions)
Ab47405, by Abcam (2 mentions)
Ab8226, by Abcam (1 mentions)
Ab243660, by Abcam (1 mentions)
5 protocols using ab155549
1
Western Blot Analysis of Protein Expression
2
IHC Quantification of SERPINE2 and CA9
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IHC was performed as previously described [26 (link)], and the primary antibodies for IHC staining included rabbit anti-SERPINE2 (AB155549, Abcam, USA) and anti-CA9 (AB243660, Abcam, USA). The IHC results were scored as H-score, including semi-quantitative grades by “IHC Profiler (Macro)” of ImageJ (version 1.53a): negative, low-positive, positive, and high-positive. The H-score = 0 * the percentage of negative cells + 1 * the percentage of low positive cells + 2 * the percentage of positive cells + 3 × the percentage of high-positive cells, which ranges from 0 to 300 [27 (link)].
3
Immunohistochemical Analysis of TRIM47 in Renal Cell Carcinoma
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After dewaxing, hydration and antigen repair, the renal cell carcinoma tissue chip was added dropwise with diluted rabbit anti-TRIM47 antibody (1:500, ab155549, Abcam, USA) and incubated overnight. The diluted anti-rabbit HRP and horseradish enzyme-labeled streptavidin working solution were added dropwise to the tissue chip the next day. The tissue chip was then stained with DAB chromogen and counterstained with hematoxylin, dehydrated and observed and counted under an optical microscope. The results of histochemical staining and the evaluation criteria of the staining intensity were scored as 0 for negative, 1 for weakly positive, 2 for moderately positive, and 3 for strongly positive. The evaluation criteria for positive cell frequency were as follows: < 5%, 0 point; 5–25%, 1 point; 26–50%, 2 points; 51–75%, 3 points; and > 75%, 4 points.
4
Immunohistochemical Analysis of TRIM47 in Renal Cell Carcinoma
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After dewaxing, hydration and antigen repair, the renal cell carcinoma tissue chip was added dropwise with diluted rabbit anti-TRIM47 antibody (1:500, ab155549, Abcam, USA) and incubated overnight. The diluted anti-rabbit HRP and horseradish enzyme-labeled streptavidin working solution were added dropwise to the tissue chip the next day. The tissue chip was then stained with DAB chromogen and counterstained with hematoxylin, dehydrated and observed and counted under an optical microscope.
The results of histochemical staining and the evaluation criteria of the staining intensity were scored as 0 for negative, 1 for weakly positive, 2 for moderately positive, and 3 for strongly positive. The evaluation criteria for positive cell frequency were as follows: <5%, 0 point; 5% -25, 1 point; 26-50%, 2 points; 51-75%, 3 points; and > 75%, 4 points.
The results of histochemical staining and the evaluation criteria of the staining intensity were scored as 0 for negative, 1 for weakly positive, 2 for moderately positive, and 3 for strongly positive. The evaluation criteria for positive cell frequency were as follows: <5%, 0 point; 5% -25, 1 point; 26-50%, 2 points; 51-75%, 3 points; and > 75%, 4 points.
5
Western Blot Analysis of Protein Markers
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Total protein was rst extracted using SDS-PAGE and then transferred to a PVDF membrane (Thermo, USA). Afterwards, the PVDF membrane was incubated with 1:1000 dilution of antibody: TRIM47 (ab155549, Abcam, USA), BAP1 (ab199396, Abcam, USA), P21 (ab188224, Abcam, USA), P53 (ab131442, Abcam, USA), SRC (ab47405, Abcam, USA), MET (ab51067, Abcam, USA), and c-Myc (ab39688,Abcam, USA). The membrane was washed and incubated with a 1:2000 dilution of horseradish peroxidaseconjugated goat anti-rabbit (Santa Cruz, USA). SuperSignalMT West Puico PLUS (Termo Scienti c, USA) was used to develop the blot, using b-actin (ab8226, Abcam, USA) as the loading control. All experiments were performed in triplicate.
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