Anti mouse cd31 rabbit polyclonal antibody

Histological sections were prepared by standard procedures and stained with hematoxylin and eosin or immunostained for hypoxic tissue, blood vessels, or collagen-I. Pimonidazole [1-[(2-hydroxy-3-piperidinyl)-propyl]-2-nitroimidazole], injected as described earlier [47 (link)], was used as a marker of tumor hypoxia, and CD31 was used as a marker of blood vessel endothelial cells. An anti-pimonidazole rabbit polyclonal antibody (Professor James A. Raleigh, University of North Carolina, Chapel Hill, NC, USA), an anti-mouse CD31 rabbit polyclonal antibody (Abcam, Cambridge, UK), or an anti-collagen-I rabbit polyclonal antibody (Abcam) was used as primary antibody. Quantitative studies were carried out on preparations cut through the central regions of tumors, and three sections of each staining were analyzed for each tumor. Microvessels were scored as described by Weidner [28 (link)]. Fraction of pimonidazole-positive tissue and fraction of collagen-I-positive tissue were assessed by image analysis [48 (link)] and were defined as the area fractions of the non-necrotic tissue showing positive staining.

Histological sections, prepared by standard procedures, were stained with hematoxylin and eosin (HE) or immunostained for blood vessels or hypoxia. CD31 was used as a marker of endothelial cells, and pimonidazole [1-[(2-hydroxy-3-piperidinyl)-propyl]-2-nitroimidazole] was used as a marker of tumor hypoxia [23 (link)]. An anti-mouse CD31 rabbit polyclonal antibody (Abcam, Cambridge, UK) or an anti-pimonidazole rabbit polyclonal antibody (Professor Raleigh, University of North Carolina, Chapel Hill, NC, USA) served as primary antibody. Diaminobenzidine was utilized as chromogen, and hematoxylin was used for counterstaining. For each immunostaining, three sections cut through central regions of every tumor were analyzed. Microvessels were identified and scored manually as number of vessels per mm² of non-necrotic tissue [24 (link)]. The hypoxic fraction, defined as the area fraction of viable tissue staining positive for pimonidazole, was determined by image analysis [25 (link)]. For each of the four PDX models, MVD was assessed in 20 early generation tumors, whereas hypoxia was quantified in 12–28 early and late generation tumors.

Histological sections were prepared by standard procedures and stained with hematoxylin and eosin (HE) or immunostained for hypoxia, blood vessels, or lymphatics. Pimonidazole [1-[(2-hydroxy-3-piperidinyl)-propyl]-2-nitroimidazole], injected as described earlier [78 (link)], was used as a marker of tumor hypoxia, and CD31 and LYVE-1 were used as markers of blood and lymph vessel endothelial cells, respectively. An anti-pimonidazole rabbit polyclonal antibody (Professor James A. Raleigh, University of North Carolina, Chapel Hill, NC, USA), an anti-mouse CD31 rabbit polyclonal antibody (Abcam, Cambridge, UK), or an anti-mouse LYVE-1 rabbit polyclonal antibody (Abcam) was used as primary antibody. Quantitative studies were carried out on preparations cut through the central regions of tumors, and three sections of each staining were analyzed for each tumor. Blood vessel density was scored by counting CD31-positive vessels in whole tumor cross-sections as described elsewhere [54 (link)]. The density of peritumoral lymphatics was assessed by counting LYVE-1-positive vessels in the muscle tissue located within a distance of 0.5 mm from the tumor surface. Fraction of pimonidazole-positive tissue was assessed by image analysis [44 (link)] and was defined as the area fraction of the viable tissue showing positive staining.