Yes, we have use it about 3years. It is fine and good results when we use it. We use about 3 microliter for 2% gel in 30 ml.

Hi Carlos, the reason you are getting the undefined bands is not because of GelRad. GelRad is simply a stain very similar to ethidium bromide and it does not matter that you are adding before the running or after.

GelRed affects the running of the DNA since it binds to the DNA. Since it affects all DNA molecules in the same way, it doesn't matter. Undefined bands usually come from the PCR reaction itself, either when it could be optimized or when the DNA template is not good enough.

In my experience (although I never used GelRed; ethidium-bromide instead, but they both employ the same interchelating principle) I've obtained better results if EtBr was added before the electrophoresis. However, mind to always add a staining agent into a gel AFTER you allow the gel to cool down to 50-60 oC, altough the producers of GelRed (see the attachment) recommend to add it in hot gel...

Thank you for the answers. The curious is that when we stain with EtBr after running, the same PCR products became ok!

Hi Carlos, I remember that a few years ago, when the lab where I was changed from ethidium bromide to SYBR safe or GelRed, there was the discussion whether these were better or not. We were always incorporating the stainings in the gel prior running, and if I recall correctly, we came to the conclusion that SYBR safe gives a weaker signal for small bands (actually it is quite difficult to see bands below 300bp if you run the gel for a long extent of the gel), while GelRed was the opposite. Some people still think that ethidium is the best and more sensitive of them all, if we exclude it`s toxicity. Nevertheless, as it was already suggested, add the stain only when you can actually hold the flask with the melted agarose with your gloved hand (this is our sensor for not too hot agarose), and before it starts to polymerise. We use 5ul of SYBR safe for 100ml of 1% agarose. I do not know however if any of this helps in your case.

In my opinion, GelRed affects migration since it binds to the DNA. In theory, it must affect all the samples in the same way.... if you have a well optimized PCR and you know the size very well, or you only want to know presence/absence of amplification it works good!