REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Mouse anti-FLAG M2	Sigma-Aldrich	F3165
Goat anti-Mouse IgG(H+L), Human ads-Alexa Fluor 488	SouthernBiotech	1031-30
Goat anti-Human IgG Fc-Alexa Fluor 647	SouthernBiotech	2048-31
Goat anti-Mouse IgG H&L (IRDYe 680RD)	abcam	ab216776
Beta Tubulin Loading Control Monoclonal Antibody (BT7R)	Thermo Fisher	MA5-16308
Bacterial strains
NEB 5-alpha Competent E. coli (High Efficiency)	NEB	C2987H
Mix & Go Competent Cells - Strain Zymo 10B	Zymo Research	T3019
Rosetta (DE3) Competent Cells	Sigma-Aldrich	70954
Chemicals, peptides, and recombinant proteins
Superior Broth	AthenaES	0105
Expi293 Expression Medium	Sigma-Aldrich	A1435101
DMEM, high glucose, pyruvate	GIBCO	11995065
Fetal Bovine Serum	GIBCO	26140079
Opti-MEM I Reduced Serum Medium, GlutaMAX Supplement	GIBCO	51985091
Penicillin-Streptomycin (5,000 U/mL)	Thermo Fisher	15070063
Lipofectamine 3000 Transfection Reagent	Thermo Fisher	L3000015
Bovine Serum Albumin	Sigma-Aldrich	A3294
EZCut TEV Protease, Recombinant	BioVision	7847-100
ACE2 Fc Chimera, Human	Genscript	Z03484
Hoechst stain	Thermo Fisher	62249
Odyssey Blocking Buffer (PBS)	LI-COR	927-40000
Surfactant P20	Cytiva	BR100054
cOmplete, EDTA-free Protease Inhibitor Cocktail	Sigma-Aldrich	11873580001
Nano-W (Methylamine Tungstate)	Nanoprobes	#2018
Commercial kits and enzymes
NEBuilder HiFi DNA Assembly Master Mix	NEB	E2621S
Esp3I	NEB	R0734
T7 DNA Ligase	NEB	M0318S
AarI (2 U/μL)	Thermo Fisher	ER1582
T4 DNA Ligase	NEB	B0202S
Promega Wizard SV 96 Plasmid DNA Purification Kit	Promega	A2250
Expi293 Transfection kit	Sigma-Aldrich	L3287
Mycoplasma Detection Kit	SouthernBiotech	13100-01
Pierce HRV 3C Protease Solution Kit	Thermo Fisher	88946
Bright-Glo Luciferase Assay System	Promega	E2610
qPCR Lentivirus Titer Kit	Applied Biological Materials (abm)	LV900
Deposited data
Raw images	This paper; Mendeley Data	https://doi.org/10.17632/k2kbt5gs6y.1
Spike Display data	This paper	Data S1
Experimental models: Cell lines
HEK293T	ATCC	CRL-3216
Expi293F Cells	Thermo Fisher	A14527
Oligonucleotides
Synthetic DNA sequences for cloning, see Table S3	Integrated DNA Technlologies	N/A
Recombinant DNA
SARS-CoV-2 S HexaPro	Hsieh et al., 2020	Addgene #154754
pcDNA5/FRT/TO/intron/GFP	Shelton et al., 2018	Addgene #113547
pYTK001	Lee et al., 2015	Addgene #65108
HDM-Hgpm2	BEI Resources	NR-52516
pRC-CMV-Rev1b	BEI Resources	NR-52519
HDM-tat1b	BEI Resources	NR-5251
Vector pHAGE2 Containing the ZsGreen Gene	BEI Resources	NR-52520
pCMV-VSV-G Envelope Vector	Cell Biolabs	RV-110
Software and algorithms
FlowJo v9	BD	N/A
NIS-Elements D	Nikon	MQS33000
cryoSPARC v3.1.0	cryoSPARC	N/A
CisTEM	https://cistem.org/software	N/A
SA3800 Spectral Cell Analyzer Software	SONY	N/A
R Tidyverse	https://www.tidyverse.org/	N/A
ggplot2 (v3.3.5)	https://ggplot2.tidyverse.org/	N/A
ChimeraX 1.1	UCSF	N/A
GraphPad Prism	GraphPad	N/A
Octet Data Analysis software (v11.1)	FortéBio	N/A
Public datasets
Expression summary of variants with single substitutions	Hsieh et al., 2020	Table S1
Yeast display RBD expression, ACE2 binding, and mAb escape data	Starr et al., 2020, 2021	https://jbloomlab.github.io/SARS-CoV-2-RBD_MAP_clinical_Abs/
Other
384-well Echo Source Plate	Biolab	PP-0200
96-well Clear Round Bottom 2.2 mL Polypropylene Deep Well Plate (Sterile)	Axygen	P-2ML-SQ-C-S
Nunc OmniTray Single-Well Plate	Thermo Fisher	140156
Protein G magnetic beads	Promega	G7471
Protein A Agarose	Thermo Fisher	15918014
Strep-Tactin Superflow resin	IBA	2-1206-025
Superose 6 increase 10/300	GE healthcare	GE29-0915-96
Cell Culture Dishes (10 cm)	VWR	10062-880
Cell Culture Plate (6-well)	VWR	10861-696
Cell Culture Plate (12-well)	VWR	10861-698
LUNA-II Automated Cell Counter	Logos biosystems	L40002
SA3800 Spectral Analyzer	SONY	N/A
5 mL HisTrap HP column	GE healthcare	17524802
Cell Imaging Dish, 35 × 10 mm	Eppendorf	0030740009
Ts2R-FL inverted research microscope	Nikon	MFA51010
CF160 Plan Apochromat Lambda 60x oil immersion objective	Nikon	MRD01605
C-FL AT DAPI/HOECHST/ALEXA FLUOR 350; Filter Cube	Chroma	96221
C-FL AT EGFP/FITC/CY2/ALEXA FLUOR 488; Filter Cube	Chroma	96226
C-FL AT TEXAS RED/mCHERY/ALEXA FLUOR 594; Filter Cube	Chroma	96231
SOLA SM II 365 LIGHT ENGINE	Lumencor	77060075
PCO.PANDA USB3.1 SCMOS CAMERA	PCO	77067012
Mini-PROTEAN TGX Precast Protein Gel	Bio-Rad	4561084
Trans-Blot SD. Semi-Dry Electrophoretic Transfer Cell	Bio-Rad	1703940
EM Grids	Electron Microscopy Sciences	CF400-Cu
Talos F200C TEM microscope	Thermo Fisher	N/A
Ceta 16M detector	Thermo Fisher	N/A
Octet Anti-Mouse Fc Capture (AMC) Biosensors	FortéBio	18-5088
Octet RED96e	FortéBio	N/A
IncuCyte ZOOM	Essen BioScience	N/A
QPix 420	Molecular Devices	N/A
Detailed bench protocols for Spike Display assays	This paper	Methods S1