Simple Western™

How Does Simple Western™ Work?

Simple Western™ is the only platform on the market that provides a fully automated Western blotting workflow, offering high reproducibility, sensitivity, and quantitation, in addition to time and cost savings. The manual steps of traditional Western blot are precisely controlled inside an automated capillary benchtop instrument. Simple Western uses CE-SDS to separate protein by molecular weight (SW-Size) or by isoelectric point using capillary isoelectric focusing (SW-Charge).

Once protein separation is complete, proteins are covalently immobilized to the capillary wall, and the steps of traditional western blot like washing, antibody incubation, and immunodetection are performed without user intervention. Upon completion, analyzed results automatically appear in the Compass for Simple Western™ software and quantitative data may be exported for further analysis.

Why Use Simple Western™?

As a capillary-based immunoassay, Simple Western™ automates the steps of traditional Western blots inside of a precisely controlled capillary environment and eliminates unreliable gel-to-membrane transfers. While Western blots are primarily qualitative and poorly reproducible, Simple Western™ produces fully quantitative and highly reproducible data with up to 6-log dynamic range and CVs consistently below 15%. Simple Western™ requires as little as 3 µL, only a fraction of the sample needed for analysis.

Simple Western’s Jess instrument provides flexible multiplex detection strategies in chemiluminescence and fluorescence channels, and the Stellar™ NIR/IR Modules provide industry-leading detection sensitivity that is 100 times more sensitive than traditional fluorescence Western blots. Simple Western provides simultaneous total protein detection, eliminating the use of unreliable housekeeping proteins for data normalization and allowing for more accurate protein expression measurements.

Simple Western™’s RePlex™ feature performs sequential immunoassay or total protein detection in the same capillary, eliminating the strip and re-probe method of traditional Western blotting. Because samples are covalently immobilized to the capillary wall, RePlex completely and reproducibly removes antibodies between probing cycles. As a result, RePlex™ assays are not impacted by signal interference or sample loss between probing cycles.

Simple Western™ covalently immobilizes samples – even within a complex matrix, – so Simple Western™ reduces matrix effects that can occur with ELISA. Following immobilization, Simple Western™ flushes the sample matrix from the capillary before immunodetection in an optimized buffer environment, further reducing matrix effects.

Simple Western™ only needs one target-specific antibody to give specific detection, which simplifies assay development relative to custom sandwich ELISA methods. Protein separation profiles on Simple Western™ allow you to identify protein isoforms or off-target antibody cross-reactivity.