4 |
Low metabolite signal |
Analyte loss due to neuron damage during isolation or culturing because of a) excessive enzymatic digestion; b) cell membrane integrity impacted by mechanical or chemical treatment |
Optimize experimental conditions of cell isolation and culturing for the sample under investigation; use stabilization solutions |
18 |
Unstable ES in ESI- MS mode. MS signal or spray current exhibits regions of
low-to-no signal
spikes, or
chaotic fluctuation
|
The ES Taylor-cone is destabilized by
bubbles in sheath flow supply (confirm via optical inspection),
electrical circuit breakdown, or
multijet/nonaxial conejet mode established (confirm via optical inspection)
|
Inspect/replace sheath flow connections
In order of preference: increase ES emitter-sampling plate distance, decrease ambient humidity, decrease sampling plate potential, or clean/polish sampling plate and ES emitter tip
In addition to b), inspect for perpendicular ES emitter-sampling plate alignment
|
19 |
Unstable ES in CE- ESI-MS mode. MS signal or spray current has regions of
low-to-no signal,
spikes (e.g., spray current reaching µA level), or
chaotic fluctuation
|
The ES Taylor-cone is destabilized by
electrolysis/solvent heating, producing bubbles adjacent to separation capillary (confirm under microscope),
electric breakdown at emitter-MS inlet region (electric sparks/arcs may are also audible), or
multijet/nonaxial conejet mode established (confirm under microscope)
|
In order of preference: Flush system and restart experiment, minimize/prevent electrolysis by lowering the CE voltage, decreasing ambient temperature, increasing sheath flow rate, or modifying the electrolyte composition
increase ES emitter-to-sampling plate, decrease CE voltage or spray voltage, rinse ES emitter thoroughly, or clean sampling plate
In addition to b), follow Troubleshooting advices given for 18
|
23 |
Separation capillary inlet end is damaged |
Capillary inlet end has been forced against the sample-loading vial and chips off or breaks (confirm under microscope) |
Avoid bending the capillary during sample loading and separation. Cleave, burn, and clean inlet end of the capillary (Step 15). Alternatively, install a new separation capillary into the platform. Measure capillary length. Note that shortening the capillary will inherently shorten the migration time of analytes, alter separation efficiency, and increase CE current. |
25 |
Electric sparks audible from the
CE platform, or
ES emitter
|
Electrical circuit shorting due to
liquid spillage or salt deposits around CE platform
salt deposits on emitter
|
Lower ambient humidity, or
inspect connections; dry components and place a drying agent (e.g., calcium sulfate) in CE enclosure; clean platform surfaces and separation capillary with isopropanol and allow to air dry
increase ES emitter-to-sampling plate distance; Sigma water- then methanol-rinse and air dry ES emitter and MS sampling plate; if problem persists, polish sampling plate.
|
28 |
Poor analytical performance observed for
test solution
single-cell extract
|
Improper performance in
sample loading, CE separation, ion generation, or MS detection
sample preparation
|
Clean system (Step 15). If problem persists, optimize ion generation and mass spectrometric analysis for the test solution (Step 18) as well as separation by analyzing decreasing amounts of analytes
Validate system performance via Troubleshooting advice 28a. Analyze rinse solution, and if positive for the compound of interest, eliminate, or minimize the duration of the water rinse (Step 4A, iv). Optimize sample preparation including the volume and composition of the extraction solution, increase sample loading volume for analysis
|