#brdu

Beyond BrdU, New Generation Cell Proliferation Imaging Analysis Kit (EdU Method)

Common methods of cell proliferation mainly include MTT method, WST-1 method and CCK-8 method, which are mainly based on the activity of cells, thus reflecting the overall proliferation effect, but unable to detect individual proliferating cells. Recognized as the most accurate method to detect cell proliferation is to directly detect DNA synthesis in cells. Previously, BrdU method was the most commonly used method. However, the method requires DNA denaturation (such as acid denaturation, thermal denaturation or DNase digestion) to expose BrdU, thus binding to BrdU antibody. The whole experiment has many influencing factors and poor stability.

Abbkine innovates and develops EdU method based on EdU incorporation and subsequent click reaction to solve this application difficulty:

Product name Cat# Fluorescent characteristic Cell Proliferation EdU Image Kit (Green Fluorescence) KTA2030 AbFluor 488 Ex/Em = 501/525 nm Cell Proliferation EdU Image Kit (Orange Fluorescence) KTA2031 AbFluor 545 Ex/Em = 546/565 nm

Principle：EdU (5-ethynyl-2´-deoxyuridine) is a nucleoside analog of thymidine and is incorporated into DNA during active DNA synthesis. Comparing to BrdU assays, the EdU-Click Assays are not antibody based and therefore do not require DNA denaturation (typically using HCl or heat or digestion with DNase) for detection of the incorporated nucleoside. Detection is based on a click reaction, a copper-catalyzed covalent reaction between an azide and an alkyne, is complete within 30 minutes.

Kit components Advantage • EdU (10mM) • AbFluor 488 azide or AbFluor 545 azide • 10×Reaction buffer • Copper • Reducing Agent • Not antibody based  • No DNA denaturation. Can maintain cell morphology and DNA integrity • Patented AbFluor 488 and 545 azide have good light stability and quenching resistance • Optimization for fluorescence microscope

Results:

New Post has been published on https://www.abbkine.com/beyond-brdu-new-generation-cell-proliferation-imaging-analysis-kit-edu-method/

Beyond BrdU, New Generation Cell Proliferation Imaging Analysis Kit (EdU Method)

Common methods of cell proliferation mainly include MTT method, WST-1 method and CCK-8 method, which are mainly based on the activity of cells, thus reflecting the overall proliferation effect, but unable to detect individual proliferating cells. Recognized as the most accurate method to detect cell proliferation is to directly detect DNA synthesis in cells. Previously, BrdU method was the most commonly used method. However, the method requires DNA denaturation (such as acid denaturation, thermal denaturation or DNase digestion) to expose BrdU, thus binding to BrdU antibody. The whole experiment has many influencing factors and poor stability.

Abbkine innovates and develops EdU method based on EdU incorporation and subsequent click reaction to solve this application difficulty:

Product name Cat# Fluorescent characteristic Cell Proliferation EdU Image Kit (Green Fluorescence) KTA2030 AbFluor 488 Ex/Em = 501/525 nm Cell Proliferation EdU Image Kit (Orange Fluorescence) KTA2031 AbFluor 545 Ex/Em = 546/565 nm

Principle：EdU (5-ethynyl-2´-deoxyuridine) is a nucleoside analog of thymidine and is incorporated into DNA during active DNA synthesis. Comparing to BrdU assays, the EdU-Click Assays are not antibody based and therefore do not require DNA denaturation (typically using HCl or heat or digestion with DNase) for detection of the incorporated nucleoside. Detection is based on a click reaction, a copper-catalyzed covalent reaction between an azide and an alkyne, is complete within 30 minutes.

Kit components Advantage • EdU (10mM) • AbFluor 488 azide or AbFluor 545 azide • 10×Reaction buffer • Copper • Reducing Agent • Not antibody based  • No DNA denaturation. Can maintain cell morphology and DNA integrity • Patented AbFluor 488 and 545 azide have good light stability and quenching resistance • Optimization for fluorescence microscope

Results:

Interests on https://www.abbkine.com/beyond-brdu-new-generation-cell-proliferation-imaging-analysis-kit-edu-method/

Beyond BrdU, New Generation Cell Proliferation Imaging Analysis Kit (EdU Method)

Did you LOL at the title? Well? Did ya? Punk?

Sorry if you did, but this post is going to be all about how we Scientists see the world of the very, very small.

When you get given a lecture about something complicated and cool, say for example Neurogenesis (the ability for the adult brain to produce new neurons from endogenous neural stem cells, an exciting area of research in which we are only just scratching the surface) you might hear something like 'Ki-67 is a protein expressed when cells divide'. But how do we know?

One way of doing this is to use a staining technique called immunohistochemistry. The procedure seems complicated, but gives pretty amazing results and is transferable between different stains on different tissues.

The theory is this:

1) Inject the substance you want to stain into a large mammal. They will then produce antibodies against it which you can harvest by dissecting the spleen.

2) The variable region of the antibody will bind to the epitope (area of the substance to which an Ig can bind) leaving the constant region free.

3) Raise antibodies against the constant region with a fluorescent tag attached. In theory, all the substances exposed to both antibodies should now fluoresce.

Here's a diagram of an Ig molecule in case you've forgotten or are a Bio-n00b.

There are a couple of problems with this method.

Ig molecules aren't as specific as all that, and you end up with what's called background staining, which can mess up your experiments. That's why you need negative controls; they show how much of the fluorescence is caused by background.

It's really sodding expensive. You have to raise large mammals and sacrifice one per Ig. So that's essentially two per experiment. So you have to buy the ready-made antibodies in. Which jacks the price up further.

I'm currently doing some lab work involving this procedure, only we're staining for something called BrdU, which we've injected into the animals ourselves. BrdU, or bromodeoxyuradine, is a thymidine analog. When cells divide, the DNA undergoes semi-conservative replication. Nucleosides such as thymidine are used in it's construction. BrdU can be taken into the DNA molecule instead of thymidine (replacing the T of ATGC genetic thumbprint) during replication. If you break up the DNA with high-strength hydrochloric acid you can then stain for the BrdU. Any cells expressing it will have been dividing. Such as neural stem cells.

You end up with awesome pictures, such as the one below.

Exciting, no? 

DE