IHC staining was performed to evaluated the expression of MCT1 and MCT4, in both tumors and paired adjacent lung tissues in each case. In brief, the formalin-fixed paraffin-embedded tissues were cut into a series of 5 µm-thick sections. The sections were de-paraffinized, rehydrated and underwent antigen retrieval using Dako EnVisionTM FLEX Target Retrieval Solution (pH 9.0) at 95 °C for 20 minutes. Anti-SLC16A3 (MCT4) antibody (Rabbit polyclonal antibody, HPA021451, Sigma-Aldrich), and anti-MCT1 antibody (Rabbit polyclonal antibody, ab238825, Abcam) were used as primary antibodies, and were diluted at a ratio of 1:100. After 2 hours of primary antibody incubation at room temperature, incubation with a secondary antibody (PV-9000, Zhongshan Jinqiao Biotechnology Co. Ltd., Beijing, China) was performed at room temperature for 30 minutes. Finally, 3,3’-diaminobenzamine and hematoxylin were used for coloration of the immune complex and nucleus, respectively. The expression of MCT4 and MCT1 was assessed by multiplying the staining intensity score and the percentage score as described in our previous study (26 ). A final score ≥6 was defined as high expression.
Hpa021451
Manufactured by Merck Group
HPA021451 is a laboratory equipment product manufactured by Merck Group. It serves as a core component in various scientific and research applications. The detailed specifications and intended use of this product are not available at this time.
Automatically generated - may contain errors
Lab products found in correlation
Envision flex target retrieval solution, by Agilent Technologies (1 mentions)
Ab238825, by Abcam (1 mentions)
Pv 9000, by Zhongshan Jinqiao Biotechnology (1 mentions)
Scn400 slide scanner, by Leica (1 mentions)
Dab chromogen, by Agilent Technologies (1 mentions)
Hematoxylin, by Agilent Technologies (1 mentions)
2 protocols using hpa021451
1
Evaluating MCT1 and MCT4 Expression in Lung Tumors
2
Immunohistochemical Analysis of Tumor Specimens
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Tumors were fixed in 4% paraformaldehyde for 48 h at 4 °C before processing for paraffin embedding. All IHC were performed in technical duplicates on 5-µm tumor sections. After being submitted to antigen retrieval with either citrate buffer at pH 5.7 or Tris-EDTA buffer at pH 9, depending on the antibody manufacturer’s instructions, sections were incubated in BSA 5% in TBS/Triton 0.05% to block non-specific binding, then overnight at 4 °C with primary antibodies for CAIX (Novus Bio, NB100-417), CD31 (Cell Signaling, 77699), CD36 (Sigma-Aldrich, HPA002018), CPT1a (Abcam, ab234111), (GLUT1 (Proteintech, 21829-1-AP), MCT1 (Proteintech, 20139-1-AP), and MCT4 (Sigma-Aldrich, HPA021451). These primary antibodies were revealed with Envision peroxidase-conjugated anti-rabbit secondary polymer antibody and DAB chromogen (Dako-Agilent, Santa Clara, CA, USA). Sections were eventually counterstained with hematoxylin (Dako-Agilent). Stained slides were then digitalized using a SCN400 slide scanner (Leica Biosystems, Buffalo Grove, IL, USA) at 240× magnification and subjected to blind analysis to obtain the percentage of stained tissue using Visiopharm software.
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