This note should inform about the correct and recommended usage of running- and sample wash buffer in combination with the Sierra SPR system.
The Sierra SPR systems are laid out with a dedicated connection/tubing for the sample wash buffer, that will be used after every injection to clean the sample needles. The connection for the sample wash buffer is located directly at the left housing panel of the system, highlighted with the letter “S” (for sample wash buffer).
The dedicated connection/tubing for the running buffer, that will be used within the microfluidics is located either directly at the left system housing panel (SPR-16, former MASS-1) or can be found on the left housing panel of the buffer box. All running buffer connection/tubing are highlighted with the letters B1 – B4 (for buffer).
Depending on the type of system usage there are different connection recommendations for both, tubing and the individual solutions.
While performing an experiment it is recommended to have a separate bottle for running buffer and a separate bottle for sample wash buffer.
During an experiment it is recommended to connect the running buffer only to at least one of the tubing named B1 – B4 at the left system housing panel or the left housing panel of the buffer box.
Any standard buffer can be used, e.g. PBS (137 mM NaCl, 10 mM Phosphate, 2.7 mM KCl, and a pH of 7.4) or Hepes-Buffer with or without additives like cofactors or DMSO, Tween20, etc.
In addition it is also recommended to connect the sample wash buffer only to the tubing named S at the left system housing panel. Always use just ddH2O or ddH2O + Tween20 (0.005 – 0.05%).
While performing the system cleaning it is recommended to insert the running buffer tubing and sample wash buffer tubing into the same bottle containing the cleaning solution.
Table 2-1: Details on composition of cleaning solution and their storage
|Desorb 1||0.5% (w/v) sodium dodecyl sulphate (SDS)||Room temperature|
|Desorb 2||50 mM glycine, pH 9.5||4-8 °C|
|Sanitize||0.5% (v/v) sodium hypochlorite||Room temperature|
The Desorb- as well as the Sanitize Routine can be found in the R3 Control software under the “Maintenance-Section”. Each cleaning command consists of three individual cleaning steps, requiring three individual cleaning solutions.
The Desorb routine washes the system with Desorb 1, Desorb 2 and ddH2O. Each solution will be used for ~41 min. for an internal cleaning of the microfluidics. In total the routine takes ~2h.
The Sanitize routine washes the system with Sanitize, ddH2O and running buffer or ddH2O. Each solution will be used for ~41 min. for an internal cleaning of the microfluidics. In total the routine takes ~2h.
For both cleaning routines it is recommended to insert all active tubing (running buffer tubing and sample wash buffer tubing) into the same bottle for each individual cleaning solution
When putting the instrument into long-term Standby or Shutdown the following procedure is recommended.
For long-term Standby as well as Shutdown it is recommended to insert all active tubing (running buffer tubing and sample wash buffer tubing) into the same bottle for each individual cleaning solution but also while system is in long-term Standby or in Shutdown.
Rule of thumb: Make fresh buffer every day!
Options for reference surface:
Options for zero injections:
Options for Start-Up Cycles:
Table 2-2: Recommended association and dissociation times
|Type of solution sample||Association Time||Dissociation Time|
|Solution sample, activation, immobilization||180-600s||0s|
|Surface sample, capture, immobilization||180-600s||0s|
|Solution sample, blocking, immobilization||180-600s||0s|
|Solution sample, capture||10-240s||0-60s|
|Solution sample, analyte||10-600s||10-3600s|
|Solution sample, regeneration||10-180s||0-60s|
|Solution sample, common||10-60s||0-60s|
|Solution sample, solv. corr.||10-60s||0-30s|
|Strength||Type of bond|
|Weak||ph > 2.5||ph < 9||ph < 9|
|10mM glycine/HCl||10mM HEPES/NaOH||25–50% ethylene glycol||0.5–1 M NaCl|
|1–10 mM HCl||0.02% SDS|
|0.5M formic acid|
|Intermediate||ph 2 – 2.5||ph 9 – 10||ph 9 – 10|
|0.5M formic acid||10–100mM NaOH||50% ethylene glycol||1–2M MgCl2|
|10mM Glycine/HCl||10mM Glycine/NaOH||0.5–0.5% SDS||1–2M NaCl|
|Strong||ph < 2||ph > 10||ph > 10|
|1M formic acid||50–100mM NaOH||25–50% ethylene glycol||2–4M MgCl2|
|10–100mM HCl||6M guanidinechloride||0.5% SDS|
|10–50mM Glycine/HCl||1M ethanolamine|
|0.1% trifluoracetic acid|