Bio-Rad eBook - 2019 - 6

Resolved:
"Gridiron"
Resolved "Gridiron"
Instant
Dry Milk

Instant
Dry Milk

It's underappreciated, but blocking is
an important step in western blotting.
It covers nonspecific binding sites on
the membrane so your antibody binds
only to its target.

Pouches
3-3.2 OZ (90.7g)
OZ (272g)
NET WT 9.6

3-3.2 OZ (90.7g) Pouches
NET WT 9.6 OZ (272g)

It's just as well.
I've seen the background
on your blots. Keep that
stuff in the kitchen!

Hup!

3-3.
NET 2 OZ
WT (90.7g)
9.6
OZ Pou
(272 ches
g)

I
Drynstan
Mil t
k

Most commonly used blocking reagents
have advantages and disadvantages.

Powdered milk

Advantages

Disadvantages

· cheap
· readily available

· contains many proteins
· not good for phospho experiments
· can cause high background

Bovine serum · good for phosphoproteins
albumin (BSA) · good for weak signals

· expensive
· contains IgG, can cause background
· low stringency, higher background

Fish gelatin

· does not contain any
serum, so does not
cross-react with
mammalian antibodies

· contains endogenous biotin

· high stringency
· low background

· may not be suitable for
phosphoproteins
· high stringency, may lower sensitivity

Casein

Even when you have
settled on the reagent, you
should ensure you are using
the right concentration
and blocking for the
optimal amount of time.
EveryB

lot

With so many choices, how do I
decide? How long do I incubate?
You can use mine. It's
universal, so it works with
all targets and blocks in
just 5 minutes.

EveryBlot

Blockheaded!
Not sure you're using the best blocking buffer for your blot and antibodies? Take the guesswork out of the
process with a five-minute blocking buffer for ALL western blots: EveryBlot Blocking Buffer.

Bio-Rad eBook - 2019

Table of Contents for the Digital Edition of Bio-Rad eBook - 2019