In fact, the main GAlthough some of the studies mentioned looked at a nonstandard wavelength (green) apart from the conventional one (blue),In summary, Ho is a vital dye that appears to be able to bind chromatin in more than one mode, which results in different spectral properties. Author information: (1)Shin Kong Wu Ho Su Memorial Hospital, Taipei, Taiwan. These include We use cookies to help provide and enhance our service and tailor content and ads.

demonstrated that hemopoietic stem cells (HSC) could be isolated by the ability of SCs to efflux Hoechst 33342 and form a side population (SP) on FACS analysis . This normalized value can then be compared to normalized values from other tissues.


Nov-Dec 2015;29(6):661-9. No other labeling should be seen with these settings. HighWire

Collect a Z stack and make a maximum intensity projection of the stack.Open the brightness and contrast window, select the color of the centrosome signal, and maximize both brightness and contrast to create a pseudobinary image of the centrosome. Thus, the percentage of dead cells can be estimated by the percentage of cells taking up the dye. For instance, the side population in one report was shown to fail to included all HSCs as there were functional long-term HSCs in the non-SP cells (Dual-wavelength analysis of Ho-stained murine bone marrow reveals two striking aspects: the presence of more than one GIf different bone marrow cell types possess distinct chromatin conformations, we would expect to find different subpopulations with Ho staining, provided that the dye concentration is adequate.

A novel method that simultaneously monitors the low fluorescence intensity of Handbook of Immunohistochemistry and in situ Hybridization of Human Carcinomas, Volume 3Handbook of Immunohistochemistry and in Situ Hybridization of Human CarcinomasEncyclopedia of Food Sciences and Nutrition (Second Edition)From the onset of the use of flow cytometry for sex separation, using By staining intact cells with the DNA binding, vital, fluorescent dye, Pathological Assessment of Experimental Models of Stem Cell and Other Regenerative TherapiesStem Cell and Gene Therapy for Cardiovascular DiseaseScienceDirect ® is a registered trademark of Elsevier B.V. By continuing you agree to the Copyright © 2020 Elsevier B.V. or its licensors or contributors.

HighWire It should be between 0.3 and 1.3 μmCollect a Z stack and make a maximum intensity projection of the stack.Open the brightness and contrast window, select the color of the centrosome signal, and maximize both brightness and contrast to create a pseudobinary image of the centrosome.Open the histogram window and select area measurement.Select the channel in which the centrosome signal was collected and adjust the low threshold slider down slightly so that everything but the centrosome signal is masked out.Using the overlay function, draw a circle ∼3 μm diameter around the first centrosome in the image. For each tissue, calculate the average and standard deviation of the fibroblast centrosome size. These properties explain the emergence of more than one population in Ho-stained bone marrow analyzed on two wavelengths (and the presence of multiple peaks on a conventional Ho histogram). Hoechst 33342 Staining Identifies the Progenitor Side Population in NOD.Cg-PrkdcscidIL2rgtmWjl/Sz Mice Harboring Pediatric Leukemias In Vivo. A standard deviation less than 15% of the average value is indicative of adequate specimen preparation and signal collection. Within each population, emission is still proportional to DNA content; thus, it is possible to define subsets in different phases of the cell cycle.Observe the cells directly under an inverted fluorescence microscope.If desired, fix the cells after vital staining of transfected cells for further detailed analysis of morphology: If other labeling is present, go back Open the measurement window and select area measurement.Select the channel in which the centrosome signal was collected and adjust the slider down slightly so that everything but the centrosome signal is masked out.Using the overlay function, draw a circle 3 μm diameter around the centrosome.Select the area button to obtain the area of the centrosome signal inside the circle. The side population pattern is a result of both the nonlinear emission of Hoechst 33342 dye under UV excitation (Petersen et al., 2004) and the distinct ability of bone marrow cells to retain Hoechst dyes, and also likely the chromatin conformation within cell populations, which results in differential access of the dye to the DNA.

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