Journal: Scientific Reports

Article Title: Characterization of human iPSC-derived sensory neurons and their functional assessment using multi electrode array

doi: 10.1038/s41598-024-55602-8

Figure Lengend Snippet: Expression of sensory neuron related genes in hiPSC, hiPSC-derived sensory neurons and human DRG. Real-time PCR showed expression of ( a ) Peripherin, ( b ) Brn3a, ( c ) TRPV1, ( d ) TRPA1, ( e ) TRPM8, ( f ) Nav1.7, ( g ) Nav1.8, ( h ) Piezo2, ( i ) TRKA, ( j ) TRKB, ( k ) TRKC, ( l ) P2X3, ( m ) H1R, ( n ) MrgprX1, ( o ) CGRP, ( p )TAC1. The square marker, the circle marker and triangle marker indicate expression of genes in hiPSC, hiPSC-derived sensory neurons and human DRG respectively. Three different lot of hiPSC-derived sensory neurons were examined. The line marker represents the mean expression of genes in hiPSC-derived sensory neurons.

Article Snippet: Fixed samples were incubated with primary antibodies against TUBB3 (1:1000, Covance, PRB-435P), Brn3a (1:25, Millipore, MAB1585), Peripherin (1:200, Millipore, AB1530), TRPV1 (1:100, Invitrogen, PA1-748), TRPM8 (1:100, NOVUS, NBP1-97311), Nav1.7 (1:100, NOVUS, NBP2-12904), TRKA (1:80, R&D SYSTEMS, MAB1751R), TRKB (1:80, R&D SYSTEMS, MAB3971), TRKC (1:100, R&D SYSTEMS, AF373), Neurofilament 200 (1:1000, Sigma, N4142), Isolectin B4 (1:1000, Thermo Fisher Scientific, I21411) over night at 4 °C.

Techniques: Expressing, Derivative Assay, Real-time Polymerase Chain Reaction, Marker

Journal: Scientific Reports

Article Title: Characterization of human iPSC-derived sensory neurons and their functional assessment using multi electrode array

doi: 10.1038/s41598-024-55602-8

Figure Lengend Snippet: Expression of sensory neuron related proteins in hiPSC-derived sensory neurons and their morphology. The cells are stained for ( a ) TUBB3, ( b ) Peripherin, ( c ) Brn3a, ( d ) TRPV1, ( e ) TRPM8, ( f ) Nav1.7, ( g ) TRKA, ( h ) TRKB, ( i ) TRKC, ( j ) NF200, ( k ) IB4. DAPI stain of nuclei is shown in blue. ( l ) Image of iPSC-derived sensory neurons which exhibit a bipolar (red arrowhead), pseudounipolar (yellow arrowhead), or multipolar morphology (green arrowhead). Scale bar represents 50 µm.

Article Snippet: Fixed samples were incubated with primary antibodies against TUBB3 (1:1000, Covance, PRB-435P), Brn3a (1:25, Millipore, MAB1585), Peripherin (1:200, Millipore, AB1530), TRPV1 (1:100, Invitrogen, PA1-748), TRPM8 (1:100, NOVUS, NBP1-97311), Nav1.7 (1:100, NOVUS, NBP2-12904), TRKA (1:80, R&D SYSTEMS, MAB1751R), TRKB (1:80, R&D SYSTEMS, MAB3971), TRKC (1:100, R&D SYSTEMS, AF373), Neurofilament 200 (1:1000, Sigma, N4142), Isolectin B4 (1:1000, Thermo Fisher Scientific, I21411) over night at 4 °C.

Techniques: Expressing, Derivative Assay, Staining

Journal: The European Journal of Neuroscience

Article Title: Identification of amino acid residues of nerve growth factor important for neurite outgrowth in human dorsal root ganglion neurons

doi: 10.1111/ejn.14513

Figure Lengend Snippet: Activation of TrkA in U2OS‐TrkA/p75‐SHC1 cells. Total chemiluminescent signal from all wavelengths was measured using reagents and recombinant cells from DiscoverX. The luminescent signal is proportional to the amount of interactions between TrkA and SHC1 or PLCγ1 present in each well. Normalized response of the interaction between TrkA and SHC1 in U2OS‐TrkA/p75‐SHC1 cells (a), or between TrkA and PLCγ1 in U2OS‐TrkA‐PLCγ1 cells (b) after 3 hr of treatment. Five (a) or four (b) independent biological repeats with four technical replicates were conducted. (c) ELISA was used to quantify direct phosphorylation of TrkA from three independent biological repeats after normalization to 100 ng/ml wild‐type NGF. Results are mean ± SEM , significant alterations compared with wild‐type NGF were found for 100 ng/ml and 300 ng/ml R100E (**** p < .0001), 30 ng/ml R100E (** p = .0016), 10 ng/ml R100E (** p = .0031), 300 ng/ml W99A (* p = .0363), 100 ng/ml K95A/Q96A (* p = .0185) and 300 ng/ml K95A/Q96A (*** p = .0006). Wild‐type NGF (black circles ●), NGF‐R100E (red squares ), NGF‐W99A (green diamonds ) or NGF‐K95A/Q96A (blue triangles )

Article Snippet: Human Phospho‐TrkA DuoSet IC ELISA and Phospho‐ERK1 (T202/Y204)/ERK2 (T185/Y187) DuoSet IC ELISA were from R&D Systems.

Techniques: Activation Assay, Recombinant, Enzyme-linked Immunosorbent Assay

Journal: The European Journal of Neuroscience

Article Title: Identification of amino acid residues of nerve growth factor important for neurite outgrowth in human dorsal root ganglion neurons

doi: 10.1111/ejn.14513

Figure Lengend Snippet: Phospho‐ERK1/2 levels in cultured U2OS‐TrkA/p75‐SHC1 cells and PC12 cells investigated with ELISA. Analysis of pERK1/2 levels in U2OS‐TrkA cells (a) or PC12 cells (b) after NGF stimulation was performed with ELISA. There were significantly increased pERK1/2 levels for NGF‐R100E (*** p = .0005) compared with wild‐type NGF and significantly lower levels for vehicle‐treated U2OS‐TrkA/p75‐SHC1 cells (* p = .0121, a). In b, significantly lower pERK1/2 levels for vehicle‐treated PC12 cells compared with wild‐type NGF were found (* p = .0121). The results are presented as mean ± SEM from four independent biological repeats with one to twelve technical replicates

Article Snippet: Human Phospho‐TrkA DuoSet IC ELISA and Phospho‐ERK1 (T202/Y204)/ERK2 (T185/Y187) DuoSet IC ELISA were from R&D Systems.

Techniques: Cell Culture, Enzyme-linked Immunosorbent Assay