Lazy Cell Lysis
For routine procedures involving cell lysis, it’s good for the lysis to be… routine. Of course there are many good and freely available lysis buffer recipes but for convenience and reproducibility you can’t beat pre-made lysis buffers.
Focusing on lysis for protein extraction, here are some of the reagents available for fast and efficient lysis of some of the most common cell types you might be using.
Reagent for E.coli lysis: Bugbuster from Novagen
Bugbuster gently breaks open E.coli cells in 20 minutes at room temperature. The lysis procedure can be enhanced by the addition of benzonase (a nuclease) and lysozyme. But I generally find that these are not required if you stick to the recommended ratio of 5ml bugbuster/1g (wet cell weight) of E.coli. Note that standard bugbuster contains primary amines, but an alternative formulation without primary amines is available.
Reagent for extremely lazy E.coli lysis: Popculture from Novagen
Popculture is a detergent-based reagent that can be added directly to an E.coli culture to lyse the cells in the medium. It is mainly used to prepare crude extracts for affinity purification by can also be used for some screening applications.
Reagent for yeast, gram positive and gram negative bacterial lysis: Y-PER from Pierce
Y-PER allows a fast and versatile lysis of yeast (and gram pos/neg bacteria). The standard protocol takes 20 minutes to produce a crude lysate that can be used for purification, screening and many other applications. Additional protocols are available that allow the reagent to be used for applications like RNA and genomic DNA extraction.
Reagent for plant cell lysis: CelLytic P from Sigma-Aldrich
CelLytic P enables extraction of proteins from fresh or frozen leaves. It has been demonstrated on tobacco, tomato, spinach, and Arabidopsis but is not limited to these. The reagent is based on a non-ionic detergent and does not denature proteins. It also contains bicine buffer, which is preferable for many biological activities.
Reagent for insect cell lysis: I-PER from Pierce
I-PER uses a non-ionic detergent formulation to lyse adherent or suspension insect cells. The extracted protein can be used for Western blotting, assays, his-purification or ion-exhange chromatography.
Reagent for Mammalian cell lysis: ProteoJET from Fermentas
Proteoject uses mild detergents for a fast and simple extraction of native, non-denatured proteins from mammalian cultured cells or tissue samples. The procedure is carried out at room temperature and can be performed on adherent cells while they are still attached to the plate - so no scraping! The isolated proteins can be used directly in many applications, including immunoprecipitation, affinity purification, reporter gene assays and electrophoretic mobility shift assays (EMSA) and Bradford assays.
Photo: Prozac74
molecular biology
Commercial lysis buffers that are enzyme based and claim to be able to lyse a broad range of bacteria seldom work. There are just so many species out there that are tough to lyse. When they take environmental samples e.g. for metagenomics, and need to be able to lyse a wide range of species, I think most use some type of mechanical disruption?
Thanks Kurt - it’s a good point that you can never make a single reagent that will lyse a wide range of bacteria since they all have different cell wall compositions.
We do some work on environmental samples in out lab and we do indeed use mechanical methods - normally glass beads for small scale work. The protocol is quite tedious though…!