Supplementary Table S1 Pre-chemo Post-chemo No ultrasound Ultrasound   Pre-chemo Post-chemo No ultrasound Ultrasound Not-evaluable 18.2% (n=6) 1.4% (n=2) 5.4% (n=3) No tumor cells 24.2% (n=8) 12.2% (n=17) 32.1% (n=18) <50% tumor content 27.3% (n=9) 47.5% (n=66) 33.9% (n=19) ≥50% tumor content 30.3% (n=10) 38.9% (n=54) 28.6% (n=16) Supplementary Table S1 Tumor cellularity in biopsies collected with and without ultrasound guidance

Supplementary Figure S1. Q-CROC-03 study: patients eligible to participate in the study were triple negative breast cancer patients with primary tumors ≥2cm and eligible to receive neoadjuvant chemotherapy. Biopsy samples were collected before initiation of treatment and/or after treatment. Surgical specimens were collected at the time of surgery.

RNAlater and Snap Frozen Shipped on dry ice 4 core biopsies from primary breast tumor And 4 pieces of tissue collected at the time of surgery Snap Frozen Formalin RNAlater and Snap Frozen Shipped on dry ice Shipped at 4°C Standard operating procedure 14-16 Gauge JGH Central Lab Standard operating procedure Supplementary Figure S2. At the time of biopsy or surgery, two samples are immediately snap frozen in liquid nitrogen, one sample is put in 1ml RNAlater and snap frozen and one sample is put in formalin. Samples are then shipped to the Central Laboratory for further processing. Established standard operating procedures were followed.

Macrodissected area containing tumor is used for DNA/RNA isolation Supplementary Figure S3 Top panel shows a frozen block of a breast biopsy specimen embedded in optimal cutting temperature medium. The pathologists circles the area where ≥50% tumor cellularity is observed to facilitate macrodissection. Bottom panel shows the frozen block macrodissected following an established standard operating procedure, only the portion with tumor is retrieved from the tissue block and used for nucleic acid extraction. SOP Macrodissected area containing tumor is used for DNA/RNA isolation