Development of a Robust Ultrasonic-Based Sample Treatment To Unravel the Proteome of OCT-Embedded Solid Tumor Biopsies

An
effective three-step proteomics workflow is proposed to overcome
the pitfalls caused by polymers present in optimum cutting temperature
(OCT)-embedded tissue during its preparation for mass spectrometry
analysis. First, the OCT-embedded tissue biopsies are cleaned using
ethanol and water in a sequential series of ultrasonic washes in an
ultrasound bath (35 kHz ultrasonic frequency, 100% ultrasonic amplitude,
2 min of ultrasonic duty time). Second, a fast ultrasonic-assisted
extraction of proteins is done using an ultrasonic probe (30 kHz ultrasonic
frequency, 50% ultrasonic amplitude, 2 min of ultrasonic duty time,
1 mm diameter tip). Third, a rapid ultrasonic digestion of complex
proteomes is performed using a microplate horn assembly device (20
kHz ultrasonic frequency, 25% ultrasonic amplitude, 4 min of ultrasonic
duty time). As a proof of concept, the new workflow was applied to
human normal and tumor kidney biopsies including chromophobe renal
cell carcinomas (chRCCs) and renal oncocytomas (ROs). A successful
cluster of proteomics profiles was obtained comprising 511 and 172
unique proteins found in chRCC and RO samples, respectively. The new
method provides high sample throughput and comprehensive protein recovery
from OCT samples.