(a) Schematic of the electroporation setup. (b) Image of the electroporation chip [27].

Flow diagram of using the electroporation system. Step 1: Collecting embryos from a Petri dish. Step 2: Placing the embryos in the chamber of the chip. Step 3: Applying Trypan blue or a toxic solution to the chamber and applying an electric field. Step 4: After electroporation, the embryos are transferred to a Petri dish.

(a) Trypan blue concentration (µg/mL) inside the chorion for application of voltages from (5–20 V) determined from the calibration curve. (b) Calculated absorbance of Trypan blue inside the chorion of embryos after application of voltages from (5–20 V) determined from the RGB values of the images. (c) The concentration of Trypan blue inside the chorion for various conditions: Left—control embryos with no voltage application only exposed to Trypan blue dye. Middle—embryos were exposed to Trypan blue and 10 V pulses. Right—embryos exposed to Trypan blue and 20 V pulses.

AuNPs inside the chorion of the embryos after exposure for 5 h (a) with AuNPs of size 20 nm (the chorion is filled with the nanoparticles) and (b) with AuNPs of size 40 nm. A purple tint is observed around the chorion, but no particle intensity is observed inside the chorion. (c) Control embryo that was not exposed to AuNPs.

(a) Embryos exposed to 8 V and 10 nm AgNP for 15 min. (b) Embryos exposed to only AgNP for 15 min. (c) Embryos exposed to AuNPs for 15 min. (d) Embryos exposed to AuNPs for 15 min after 8V application. (e) Absorbance gradient for nanoparticles. The statistical significance for p-value < 0.05 is presented as ‘*’ in the graph.

Control experiments: (a) Absorbance calculation for concentration 20–100 µg/mL AgNPs exposure for 5 h. (b) Deformity and survival rate of embryos after exposure to 20–100 µg/mL AgNPs for 24 h.

Application of electric field: (a) Absorbance calculation for concentration of 60 µg/mL of AgNP exposure for 40 min. “*” indicates statistically significant difference (p-value < 0.05) when compared with only silver particle. (b) Deformity rate of embryos after exposure to 60 µg/mL of AgNPs for 24 h for five repetitions. (c) Average deformity rate of five repetitions after exposure to 60 µg/mL of AgNPs for 24 h. Here, Only Nano indicates the embryos were exposed to AgNPs only. Only V indicates embryos were exposed to only voltage pulses with a chamber filled with E3 media instead of nanoparticles. Nano+V indicates embryos that are exposed to both nanoparticles and voltage pulses. “*” indicates statistically significant difference (p-value < 0.05) when compared with control 1 (No V+ NO particle).

Deformities observed from exposure to 10nm AgNPs for 40 min after electroporation (a) Control with no deformity. (b) Abnormal middle body shape; (c) abnormal tail; (d) tilted tail.

Condition of heart rate for exposure to 10nm AgNPs for 40 min. “*” indicates statistically significant difference (p-value < 0.05) when compared with control 1 (No V+ NO particle).

Survival rate for exposure of 10nm AgNPs for 40 min after electroporation (N = 5).