In vivo bromodeoxyuridine incorporation in normal mouse kidney: Immunohistochemical detection and measurement of labelling indices

Robertson, Helen Lee; Wheeler, J. W.; Morley, A. R.

doi:10.1007/bf02386007

Cited by 17 publications

(10 citation statements)

References 16 publications

(14 reference statements)

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“…The proliferation rate of EGF-labeled cells was measured by administering a single injection of 50 g/g body wt of BrdU (Boehringer Mannheim, Mannheim, Germany) to P3 and P7 animals 18 h before death; the kidneys were subsequently preserved for immunohistochemistry. BrdU is a thymidine analog incorporated into DNA during the S phase of the cell cycle and can be subsequently detected in tissue sections with specific antibodies raised against it (29).…”

Section: Animals and Brdu Treatmentmentioning

confidence: 99%

Expression of epidermal growth factor in the developing rat kidney

Jung¹,

Song²,

Li³

et al. 2005

American Journal of Physiology-Renal Physiology

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Epidermal growth factor (EGF) is important in mammalian renal development. In our study, we investigated the detailed distribution and the time of the first appearance of EGF in developing rat kidney. Kidneys from embryonic 18 (E18)- and 20-day-old (E20) fetuses, postnatal 1 (P1)-, 3 (P3)-, 7 (P7)-, 14 (P14)-, and 21-day-old (P21) pups, and adults were processed for immunohistochemistry and electronmicroscopy. In adult rat kidney, EGF immunoreactivity was found in distal tubule including the thick ascending limb (TAL) and portion 1 of distal convoluted tubule (DCT1), whereas no EGF immunoreactivity was seen in portion 2 of distal convoluted tubule (DCT2) and connecting tubule. In developing kidney, EGF-positive cells first appeared at P3 and were localized in the middle portion of the differentiating TAL of the corticomedullary junction. By P7, the abundance of EGF expression had dramatically increased in the medullary TAL. Between P14 and P21, EGF immunoreactivity was found in the TAL and the DCT for the first time. However, EGF-positive and EGF-negative cells were in the TAL in developing rat kidney. EGF-positive cells did not differ from negative cells in the expression of sodium transport proteins or in the proliferation rate at P3 and P7. In the TAL, smooth-surfaced cells had strong EGF immunoreactivity, but no EGF immunoreactivity was seen in the rough-surfaced cells with well-developed microvilli. Our results suggest that the expression of EGF in developing kidney plays an important role in the regulation of growth and differentiation of the loop of Henle during kidney development and that this may act in the paracrine mode.

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Section: Animals and Brdu Treatmentmentioning

confidence: 99%

Expression of epidermal growth factor in the developing rat kidney

Jung¹,

Song²,

Li³

et al. 2005

American Journal of Physiology-Renal Physiology

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“…BrdU/antiBrdU antibody immunocytochemical staining has been used for various tissues. Procedures for cell dynamic studies have been followed with individual light and electron microscopic techniques (1,4,9,14,17,19). The cell type observed with light microscopy is not always delivering the same information observed with the electron microscope.…”

Section: Discussionmentioning

confidence: 99%

“…Resolution power at the ultrastructural level is limited by the diameter of the silver grains and the range of isotopes (1,8,9,17,19). As an alternative method to radioautography, immunoelectron microscopic staining using monoclonal antibodies has been developed for localizing the sites of DNA replication (14). For this purpose, the monoclonal antibody against bromodeoxyuridine (BrdU), an analogue of thymidine, has been used successfully (2,4,7,12,18).…”

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confidence: 99%

Immunoelectron Microscopic Labeling of Digestive Tract Stem Cells by Means of Bromodeoxyuridine Antibody.

Tsuyama

Yang

Kanae

et al. 1994

Acta Histochem. Cytochem

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As an alternative method to 3H-thymidine radioautography, immunohistochemical staining against incorporated bromodeoxyuridine (BrdU), an analogue of thymidine, was performed on rat stomach and small intestine organs embedded in Lowicryl K4M resin. After the administration of BrdU to animals intraperitoneally, organ tissues were removed and processed in a routine manner for resin embedding procedures. Semithin and ultrathin serial sections were cut and incubated in mouse anti-BrdU monoclonal antibody and stained with anti-mouse IgG or protein A coupled with gold colloid. Semithin sections were further intensified by physical development using silver acetate as an ion donor. In the electron microscopic labeling, pretreatment with hydrogen peroxide increased the labeling intensity.The findings from both light and electron microscopic observation were compared and a good correlation was obtained. Various labeling patterns were found in S-phase cell nuclei from stomach and small intestines in vivo. Labeling patterns of nuclei matrix were divided into five subtypes according to labeling patterns at the electron microscopic level.

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“…Most studies using BrdU involve rapidly proliferating tissues. However, BrdU incorporation has also been used successfully in systems exhibiting low levels of proliferation, although such studies are sparse (5,11,12,15) heir results were compatible with measurements made using TdR. On the other hand, DeFazio et al (5) claimed that normal liver remained unstained after three days of exposure to BrdU.…”

mentioning

confidence: 87%

In vivo Bromodeoxyuridine Incorporation in Normal Rat Liver: Immunohistochemical Detection of Streaming Cells and Measurement of Labelling Indices.

Arber¹,

Zajicek²,

Schapiro

et al. 1994

Acta Histochem. Cytochem

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The aim of the present study was to determine if normal liver cell kinetics can be evaluated using bromodeoxyuridene (BrdU) as the label. BrdU was injected intraperitoneally into 10 male adult rats, 5 were sacrified after 1 hr, and the remainder after 30 days. The livers were fixed in 70% ethanol, embedded in paraffin and histological slides were prepared.The labelled nuclei were revealed using indirect immunoperoxidase staining and easily counted by light microscopy.One hr after labelling, labelled hepatocytes and littoral cells were detected within 164 ± 12.3 ~tm and 161 ± 13 pin respectively from the portal tract rim. After 30 days, both cell types reached the respective distances of 290 ± 14.6 pm and 294 ± 17.7 pm. Their respective displacement velocities were 4.2 pm/d and 4.3 pm/d. These results were identical to those we obtained using tritiated thymidine.This laboratory method provides a useful and simpler alternative method to tritiated thymidine.Cell kinetics have been traditionally studied by measuring tritiated thymidine (TdR) incorporation into DNA in the S-phase of the cell cycle. The validity of BrdU labelling for cell kinetic studies has been well established in comparison with other methods and it is now regarded as a useful alternative to TdR (13). Most studies using BrdU involve rapidly proliferating tissues. However, BrdU incorporation has also been used successfully in systems exhibiting low levels of proliferation, although such studies are sparse (5,11,12,15) heir results were compatible with measurements made using TdR. On the other hand, DeFazio et al. (5) claimed that normal liver remained unstained after three days of exposure to BrdU.The aim of the present study was to demonstrate that BrdU can serve as a reliable marker for determination of liver cell kinetics in normal mature rats not exposed to any liver insult. The histochemical method used is quick and relatively simple (Darmon et al.,(4) ), and the results are comparable to those we obtained using TdR (1-3, 17). MATERIALS AND METHODSTen male adult rats, random-bred Tel Aviv University strain weighing 250-300 g, were injected intraperitoneally (IP) with 50 mg/kg body weight BrdU (Sigma Chemical Company Ltd, Poole, Dorset) at a concentration of 6 mg/ml in 0.05 M phosphate buffered saline, pH 7.4 (PBS). Five rats were sacrified after one hr, the remaining five rats were fed and watered, ad libitum, for 30 days and then sacrified. The livers, and a short section of the small intestine were immediately removed and fixed in 70% ethanol

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In vivo bromodeoxyuridine incorporation in normal mouse kidney: Immunohistochemical detection and measurement of labelling indices

Cited by 17 publications

References 16 publications

Expression of epidermal growth factor in the developing rat kidney

Expression of epidermal growth factor in the developing rat kidney

Immunoelectron Microscopic Labeling of Digestive Tract Stem Cells by Means of Bromodeoxyuridine Antibody.

In vivo Bromodeoxyuridine Incorporation in Normal Rat Liver: Immunohistochemical Detection of Streaming Cells and Measurement of Labelling Indices.

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