Pacific Blue™ Annexin V/SYTOX™ AADvanced™ Apoptosis Kit, for flow cytometry

Este producto detecta la externalización de la fosfatidilserina en células apoptóticas utilizando anexina V recombinante conjugada con la tinción violetaMás información

Número de catálogo	Cantidad
A35136	1 Kit

Número de catálogo A35136

Precio (CLP)

620.835

Añadir al carro de la compra

Cantidad:

1 Kit

Precio (CLP)

620.835

Añadir al carro de la compra

Este producto detecta la externalización de la fosfatidilserina en células apoptóticas utilizando anexina V recombinante conjugada con la tinción violeta fluorescente Pacific Blue™ y células muertas mediante la tinción SYTOX™ AADvanced™. Después de teñir una población celular con tinción Pacific Blue™, anexina V y SYTOX™ AADvanced™, las células apoptóticas muestran fluorescencia violeta, las células muertas muestran fluorescencia roja, y las vivas muestran poca o ninguna fluorescencia. Estas poblaciones se distinguen fácilmente con un citómetro de flujo con las líneas de excitación de 405 nm y 488 nm. Existe muy poca superposición espectral entre los dos colorantes, por lo tanto, se necesita muy poca compensación. Cada kit contiene suficientes reactivos para aproximadamente 50 pruebas de citometría de flujo.

Consulte la guía de selección para todos los ensayos de apoptosis para citometría de flujo.

Para uso exclusivo en investigación. No apto para uso en procedimientos diagnósticos.

Especificaciones

Excitación/emisiónPacific Blue™: 415⁄455, SYTOX™ AADvanced™: 546⁄647

Líneas láser del citómetro de flujo405, 488

Para utilizar con (aplicación)Citometría de flujo

Para utilizar con (equipo)Citómetro de flujo

N.º de reacciones50

Tipo de productoKit de apoptosis

Cantidad1 Kit

Condiciones de envíoHielo húmedo

ConjugadoColorante para células muertas Pacific Blue™, SYTOX™ AADvanced™

FormatoTubo

Unit SizeEach

Contenido y almacenamiento

Contiene 1 vial de conjugado de anexina V, Pacific Blue™ (250 µl), 1 vial de tinción de células muertas SYTOX™ AADvanced™ y 1 frasco de tampón de unión de anexina (solución 5X, 15 ml) y 1 vial de DMSO (100 µl).

Almacenar en el refrigerador (2–8 °C) y proteger de la luz.

Preguntas frecuentes

I want to study apoptosis using an Annexin V conjugate, but with adherent cells via microscopy instead of flow cytometry. Can this be done?

It has been done, but we don‘t recommend it. Both healthy cells and apoptotic cells possess phosphatidylserine on the cell surface, which can be detected with Annexin V, but apoptotic cells have significantly more of it. You can easily tell the difference between these two populations with flow cytometry, because flow cytometers are more sensitive and have a higher throughput. But with a microscope, you cannot always tell the difference, especially for adherent cells. Instead, for microscopy, we recommend a different technique, such as detecting caspases with CellEvent Caspase Detection Reagents.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

I trypsinized my adherent cells and labeled with annexin V, and now my flow data is showing a high percentage of apoptotic cells even for control, untreated cells. What is the problem?

Trypsinization or mechanical scraping of cells temporarily disrupts the plasma membrane, allowing annexin V to bind phosphatidylserine on the cytoplasmic surface of the cell membrane and thus leading to false positive staining. Allow the cells to recover for about 30 minutes in optimal cell culture conditions and medium after trypsinizing/scraping so that they can recover their membrane integrity before staining. For lightly adherent cell lines, such as HeLa and NIH 3T3, another option is to use non-enzyme treatments like Gibco Cell Dissociation Buffer (Cat. No. 13151014).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Can I detect annexin V staining in an imaging assay?

Annexin V staining is not typically used in imaging experiments; it is a better reagent for flow cytometry analysis. All cells will stain to some extent, so it can be difficult to distinguish a relatively bright annexin V-stained cell from a dimmer non-apoptotic cell. Caspase activation, detected using our CellEvent Caspase 3/7 or Image-iT LIVE Caspase detection kits, is a better method for detecting apoptosis in an imaging assay.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

When should I stain adherent cells with annexin V for flow cytometric analysis? Before or after I trypsinize them?

Trypsinize first and then allow the cells to recover about 30 minutes in optimal cell culture conditions and medium before staining with annexin V conjugates. Trypsinization or mechanical scraping of cells temporarily disrupts the plasma membrane, allowing for annexin V to bind phosphatidylserine on the cytoplasmic surface of the cell membrane and thus leading to false positive staining. For lightly adherent cell lines such as HeLa and NIH 3T3, you could use a less harsh (non-enzymatic) dissociation product like Gibco Cell Dissociation Buffer (Cat. No. 13151014).

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Can I fix my cells after annexin V staining?

Yes, this is possible. We have established protocols for annexin V staining combined with intracellular staining of lymphocytes that can be found here. The most important step is to leave some binding buffer in the suspension when fixation is started. Compared to staining of live cells, the intensity of the annexin V signal may be somewhat reduced.