Table 3.

Summary of key microfluidic intracellular delivery techniques covered in this review

				Cargo type
Mechanism	Method	Delivery principle	Cell type (primary cell)	Fluorescence probe	Functional nanomaterials/macromolecule	Throughput (cell processing rate)	Comments
Mechanoporation	Fluid shear	Shear force	DU145,[ 45 ] NIE‐115,[ 46 ] rat primary neuron [ 46 ]	Calcein,[ 45 ] FITC–dextran,[ 45 ] FITC–BSA[ 45 ]	GFP pDNA[ 46 ]	N/A	–
		Vortex shedding	Human T cell [ 48 ]	N/A	GFP mDNA[ 48 ]	107 cells min−1[ 48 ]	Under commercialization by Indee Labs[ 48 ]
		Spiral vortex	HEK293,[ 49 ] ES2,[ 49 ] 3T3,[ 49 ] KU812,[ 49 ] HeLa,[ 49 ] HCT116,[ 49 ] MCF7,[ 49 ] HDFa,[ 49 ] K562,[ 35 , 49 ] MDA231[ 35 , 49 ]	FITC–dextran[ 35 , 49 ]	DNA nanotube,[ 49 ] DNA donut,[ 49 ] GFP mRNA,[ 35 ] GFP pDNA,[ 35 , 49 ] GNP,[ 35 ] DOX‐MSNs[ 35 ]	106 cells min−1[ 35 , 49 ]	Clogging‐free operation;[ 35 , 49 ] under commercialization by MxT Biotech[ 35 ]
	Constriction	Cell squeezing	HeLa,[ 53 , 54 , 56 ] murine ESC,[ 53 , 56 ] NuFFs,[ 53 ] murine dendritic cell,[ 53 ] murine macrophage,[ 53 ] murine B cell,[ 53 , 55 ] murine T cell,[ 53 ] human T cell,[ 57 ] Jurkat,[ 62 ] K562,[ 62 ] HEK293t[ 62 ]	FL‐dextran,[ 53 , 55 , 56 , 57 , 62 ] FL‐protein[ 53 , 62 ]	siRNA,[ 53 ] GFP pDNA,[ 56 ] CNT,[ 53 ] GNP,[ 53 ] Qdot,[ 54 ] antigen,[ 55 ] Cas9‐RNP[ 57 ]	106 cells min−1[ 53 , 54 , 56 , 57 ]	Under commercialization by SQZ Biotech (in Phase I trial; NCT04084951)[ 53 ]
		Deformation array	HEK293t,[ 58 ] MCF7,[ 58 ] SU‐DHL‐1,[ 58 ] ES AB2.2,[ 58 ] PC‐3,[ 58 ] SUM159,[ 58 , 59 ] HeLa,[ 58 ] HL60,[ 59 ] SK‐BR‐3,[ 59 ] human T cell,[ 59 ] MDA231,[ 58 , 60 ] NIH 3T3,[ 61 ] human ASC [ 61 ]	FITC–ssDNA,[ 58 ] FL‐dextran,[ 59 , 60 , 61 ] FITC–siRNA[ 59 , 60 ]	siRNA,[ 58 , 60 ] GFP pDNA,[ 58 , 61 ] CRISPR pDNA,[ 58 ] Cas9 RNP[ 58 , 59 ]	105 cells min−1[ 61 ]	–
		Volume exchange	PC3,[ 33 ] PBMC,[ 33 ] K562,[ 33 , 34 ] HL60,[ 34 ] HEY,[ 34 ] HEK293,[ 34 ] OVCAR‐3[ 34 ]	FITC–dextran,[ 33 , 34 ] RNA probe[ 33 ]	GFP mRNA,[ 33 ] GFP pDNA,[ 33 ] nanobead[ 33 ]	106 cells min−1[ 34 ]	Under commercialization by CellFE[ 33 ]
	Poking	Cell poking	MDA231,[ 75 ] HSC,[ 75 ] NIH3T3,[ 76 ] HEK293,[ 76 ] MDCK,[ 76 ] HeLa,[ 76 ] HCT116[ 76 ]	FL‐dextran,[ 75 , 76 ] Alexa488‐protein[ 76 ]	RFP pDNA,[ 75 ] Cas9 RNP,[ 75 ] siRNA[ 76 ]	103 cells min−1,[ 75 ] 105 cells min−1[ 76 ]	–
		Static poking	Jurkat,[ 77 ] K562,[ 77 ] human T cell [ 77 ]	PI dye[ 77 ]	GFP pDNA[ 77 ]	104 cells per run[ 77 ]	Under commercialization by Basilard Biotech[ 77 ]
		Hydrodynamic cell stretching	A2780cis,[ 78 ] ES2,[ 78 ] HEK293,[ 78 ] MDA231,[ 15 , 78 ] K562,[ 15 , 78 ] HEK293t,[ 15 ] 3T3,[ 15 ] DC 2.4,[ 15 ] HeLa,[ 15 ] human MSC,[ 15 ] human ADSC,[ 15 ] murine BMDC [ 15 ]	FITC–dextran[ 15 , 78 ]	DNA nanostructure,[ 78 ] CRISPR/Cas9,[ 78 ] siRNA,[ 15 , 78 ] GFP pDNA,[ 15 , 78 ] GFP mRNA,[ 15 ] Qdot,[ 15 ] nanobead[ 15 ]	106 cells min−1[ 15 , 78 ]	Demonstrated 300 nm nanoparticle delivery;[ 15 ] under commercialization by MxT Biotech[ 15 ]
	Microinjection	Microinjection on chip	HeLa,[ 84 , 90 ] zebrafish embryo,[ 83 ] Drosophila embryo[ 91 , 93 ]	FL‐dextran,[ 84 , 90 ] methylene blue,[ 83 ] rhodamine B,[ 83 , 93 ] fluorescein,[ 91 ] NaN3 [ 93 ]	siRNA[ 91 ]	60 cells min−1[ 84 ]	–
		Probe tip injection	C2C12,[ 186 ] NG108,[ 186 ] MCF7,[ 187 ] RAW 264.7,[ 187 ] RKO[ 187 ]	FITC‐sodium salt,[ 186 ] CellTracker green[ 186 ]	FITC‐nanodiamond[ 187 ]	N/A	Commercialized by Cytosurge AG[ 186 ]
	Acoustic cavitation	Ultrasonic wave	H9C2[ 111 ]	FITC‐dextran[ 111 ]	Pharmaceutical agents (doxorubicin, apigenin, and luteolin)[ 111 ]	N/A	No microbubble required[ 111 ]
		Bubble oscillation	HUVEC,[ 114 ] primary rat aortic‐smooth muscle cell [ 99 ]	DOX‐liposome,[ 114 ] calcein[ 99 ]	N/A	N/A	Vessels‐on‐a‐chip cytotoxic drug delivery[ 114 ]
		Surface acoustic wave	MCF7[ 115 ]	PI dye[ 115 ]	N/A	N/A	–
		Acoustic wave cell manipulation	Jurkat,[ 117 ] MEF,[ 117 ] PBMC,[ 117 ] HSPC [ 117 ]	Cy3‐labeled DNA[ 117 ]	GFP pDNA[ 117 ]	105 cells min−1[ 117 ]	Not requiring microbubble[ 117 ]
	Laser cavitation	Single bubble cavitation	HL60,[ 118 ] myeloma cell[ 119 ]	Trypan blue[ 118 , 119 ]	N/A	N/A	–
		Microbubble array cavitation	HeLa,[ 120 ] PB‐MDMs,[ 120 ] NHDFs,[ 120 ] RPTECs [ 120 ]	FITC–dextran[ 120 ]	Antibody,[ 120 ] enzyme,[ 120 ] bacteria,[ 120 ] nanobead[ 120 ]	105 cells min−1[ 120 ]	Macromolecule delivery[ 120 ]
Electroporation	Cell trapping‐based electroporation	Microhole trapping	ND‐1[ 149 , 150 ]	YOYO‐1 dye[ 150 ]	EGFP pDNA[ 150 ]	N/A	–
		Channel trapping	HeLa,[ 151 , 152 , 153 ] C2C12,[ 155 ] human MSC [ 155 ]	Trypan blue,[ 151 ] calcein,[ 151 , 152 , 153 ] FL‐dextran,[ 152 , 153 ] PI dye[ 155 ]	EGFP‐ERK1 pDNA[ 155 ]	N/A	–
		Membrane sandwich electroporation	NIH 3T3,[ 156 ] mouse ES [ 157 , 158 ]	N/A	GFP pDNA,[ 156 , 157 ] SEAP pDNA[ 156 , 157 , 158 ]	104 cells per run[ 157 ]	–
	Nanochannel electroporation	Single‐cell electroporation	Jurkat,[ 163 ] K562,[ 163 ] mouse embryonic fibroblasts [ 163 ]	PI dye,[ 163 ] Cy3‐ODN[ 163 ]	GAPDH molecular beacon,[ 163 ] siRNA,[ 163 ] Qdot,[ 163 ] Cy3‐labeled GFP pDNA[ 163 ]	N/A	Faster delivery compared with bulk and microelectroporation[ 163 ]
		Nanochannel array electroporation	NK‐92,[ 166 ] H9C2,[ 166 , 167 ] MEF,[ 165 , 167 ] mouse cardiomyocyte,[ 165 ] MDA231,[ 168 ] CHO,[ 168 ] HT1080[ 168 ]	PI dye,[ 166 , 167 , 168 ] FAM‐ODN,[ 167 ] Co2+,[ 168 ] Alexa488‐BSA[ 168 ]	CAR/GFP pDNA,[ 166 ] GFP pDNA,[ 165 ] OSKM pDNA,[ 167 ] miRNA,[ 165 ] mCherry pDNA[ 168 ]	104 cells per cm2,[ 167 ] 106 cells per run[ 169 ]	mRNA internalization into exosome[ 169 ]
		Nanopore electroporation	HeLa,[ 170 ] HEK293,[ 170 ] 3T3,[ 170 ] Jurkat[ 170 ]	N/A	mCherry mRNA,[ 170 ] GFP pDNA[ 170 ]	104 cells per run[ 170 ]	–
		Nanoinjection	Human MSC [ 171 ]	RFP[ 171 ]	GFP pDNA[ 171 ]	N/A	Under commercialization by Femtobiomed[ 171 ]
		Nanostraw electroporation	CHO,[ 174 ] HEK293t,[ 174 ] HEK293,[ 176 ] hiPSC‐CMs,[ 176 ] HSCs,[ 176 ] HFs,[ 176 ] mouse glia,[ 176 ] mouse neuron,[ 176 ] HSPCs [ 142 ]	PI dye,[ 174 ] FL‐dextran,[ 142 ] FITC‐OGN[ 142 ]	RFP pDNA,[ 174 ] mCherry mRNA,[ 176 ] GFP mRNA,[ 142 , 176 ] STIM1 protein,[ 176 ] Cas9 RNP,[ 176 ] siRNA[ 142 ]	105 cells per run[ 176 ]	Under commercialization by NAVAN Technologies[ 174 ]
	Nanochannel electroporation	Nanofountain electroporation	HeLa,[ 181 , 182 ] HT1080,[ 182 ] HEK293[ 183 ]	FL‐dextran,[ 181 , 182 ] BSA[ 181 , 182 , 183 ]	GAPDH‐molecular beacon,[ 181 , 182 ] GFP pDNA,[ 181 , 183 ] GFP knockdown CRISPR/Cas9[ 183 ]	N/A	–
	Flow‐through electroporation	Flow‐through DC electroporation	CHO,[ 189 , 190 , 191 , 196 ] K562[ 197 ]	SYTOX dye,[ 189 , 190 ] PI dye[ 197 ]	GFP pDNA,[ 190 , 191 , 197 ] Qdot[ 196 ]	105 cells min−1,[ 197 ] 106 cells min−1,[ 190 ] 107 cells min−1[ 191 ]	–
		Flow‐through AC electroporation	Huh‐7,[ 188 ] K562,[ 198 ] CT26.WT,[ 198 ] C3H10T1/2,[ 199 ] HeLa,[ 200 ] CHO,[ 193 ] HEK293a,[ 201 ] Neuro‐2a,[ 201 ] HUVEC,[ 201 ] HSF [ 201 ]	PI dye,[ 198 ] FL‐dextran,[ 200 ] phallotoxin[ 200 ]	β‐galactosidase pDNA,[ 188 ] GFP pDNA,[ 188 , 193 , 198 , 199 , 201 ] GFP siRNA[ 200 ]	103 cells min−1,[ 199 ] 104 cells min−1,[ 201 ] 106 cells min−1[ 198 ]	–
		Flow electroporation with laminar flow	Yeast cell,[ 202 ] HEK293a,[ 203 ] HEK293,[ 203 ] HeLa,[ 203 ] C2C12,[ 203 ] Neuro‐2a,[ 203 ] PC12,[ 203 ] human T cell [ 205 , 206 ]	Fluorescein[ 202 ]	siRNA,[ 203 ] GFP pDNA,[ 203 ] mCherry mRNA[ 205 , 206 ]	104 cells min−1,[ 202 ] 105 cells min−1,[ 203 , 205 ] 107 cells min−1[ 206 ]	Electroporation assisted by acoustophoretic cell manipulation[ 205 ]
		Flow electroporation with Dean/vortex flow	CHO,[ 204 ] MDA231,[ 207 , 208 , 210 ] HEK293,[ 208 ] MCF7,[ 208 ] HeLa[ 208 ]	YOYO‐1 dye,[ 204 ] PI dye,[ 208 , 210 ] FL‐protein[ 208 ]	GFP pDNA,[ 204 , 210 ] RFP pDNA,[ 210 ] drug (gemcitabine, bleomycin, topotecan, and quercetin),[ 207 ] siRNA,[ 208 ] miRNA[ 208 ]	105 cells min−1[ 204 ]	Cytotoxic drug delivery[ 207 ]
	Droplet electroporation	Flow‐through droplet electroporation	Yeast cell,[ 212 ] CHO,[ 213 ] green microalgae (C. reinhardtii)[ 214 ]	Fluorescein,[ 212 ] SYTOX green dye[ 213 ]	GFP pDNA,[ 213 ] DNA fragment[ 214 ]	104 cells min−1[ 213 ]	Delivery assisted by Dean vortices[ 214 ]
		EWOD droplet electroporation	E. coli [ 216 ]	N/A	Antibiotic resistance pDNA[ 216 ]	N/A	Transfection efficiency assessed using colony forming unit[ 216 ]