Systems for performing two-dimensional chromatography are disclosed. The system includes a first switching valve fluidly coupled to a first separation column. The system includes a second switching valve coupled to a second separation column. The second switching valve is fluidly coupled to the first switching valve. The system includes an input/output valve in fluid communication with the first switching valve and the second switching valve that is configured to direct one or more mobile phases to the first switching valve. The system includes a sampling valve fluidly coupled to the first switching valve that includes at least one fraction capturing device. Methods of separating compounds using the systems are disclosed. Methods of retrofitting existing two-dimensional chromatography systems are also disclosed.

The present application relates to systems and methods for assaying presence of large molecule analytes, such as proteins, e.g., antibodies, antigens, receptors, and the like, using a targeted two-dimensional liquid chromatography, tandem mass spectrometry (2D-LC-MS/MS) system, optionally combined with affinity capture. In some aspects, the system is partially or fully automated. In some aspects, the system may allow detection of protein biomarkers (e.g., antibodies or antigens) from clinical or nonclinical biological tissue or fluid samples in the pg/mL to ng/mL range.

A fluid processing apparatus includes a first fluid separation apparatus including a first fluid pump, configured for driving a first mobile phase, and a first separation unit configured for separating a fluidic sample when within the first mobile phase. A sampling unit includes a modulation buffering unit and a modulation drive, wherein the modulation drive is configured for introducing fluid into the modulation buffering unit. A switching unit is configured, in a first switching state, for introducing fluid into the modulation buffering unit from downstream of the first separation unit, and, in a second switching state, for introducing fluid buffered in the modulation buffering unit in a first flow path between the first fluid pump and the first separation unit.