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STEMCELL Technologies Inc easyseptm direct human neutrophil isolation kit
Easyseptm Direct Human Neutrophil Isolation Kit, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/direct+human+neutrophil+isolation+kit/easysep+human+t+cell+isolation+kit/pm40664875-183-24-37
Average 90 stars, based on 1 article reviews
easyseptm direct human neutrophil isolation kit - by Bioz Stars, 2026-09
90/100 stars

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Related Articles

Incubation:

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection.
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37 ̊C the leucocytes were washed (500xg, 10 min, 4 ̊C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37°C the leucocytes were washed (500xg, 10 min, 4°C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Magnetic Cell Separation:

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection.
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37 ̊C the leucocytes were washed (500xg, 10 min, 4 ̊C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37°C the leucocytes were washed (500xg, 10 min, 4°C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Sterility:

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection.
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37 ̊C the leucocytes were washed (500xg, 10 min, 4 ̊C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37°C the leucocytes were washed (500xg, 10 min, 4°C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Purification:

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection.
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37 ̊C the leucocytes were washed (500xg, 10 min, 4 ̊C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37°C the leucocytes were washed (500xg, 10 min, 4°C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Isolation:

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection.
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37 ̊C the leucocytes were washed (500xg, 10 min, 4 ̊C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: β 2 -integrin LFA1 mediates airway damage following neutrophil transepithelial migration during respiratory syncytial virus infection
Article Snippet: .. Purify neutrophils using a direct human neutrophil isolation kit (STEMCELL), as per the manufacturer’s instructions. ..

Article Title: Panton-Valentine leukocidin-induced neutrophil extracellular traps lack antimicrobial activity and are readily induced in patients with recurrent PVL+-Staphylococcus aureus infections.
Article Snippet: .. Human neutrophils were isolated by layering whole blood over Histopaque-1119 (Sigma-Aldrich) followed by a discontinuous Percoll gradient (Amersham Biosciences) as previously described.46 Alternatively, they were also isolated using the direct human neutrophil isolation kit (EasySep; STEMCELL Technologies) following manufacturer’s instructions. .. All experiments were performed in Seahorse XF RPMI medium (Agilent) supplemented with 2 mM glutamine, 10 mM HEPES, 1 mM glucose, and 0.1% human serum albumin (HSA) at pH 7.4, except where mentioned.

Article Title: Mucosal Therapy of Multi-Drug Resistant Tuberculosis With IgA and Interferon-γ.
Article Snippet: Neutrophils were purified from blood using HetaSep (Stemcell, Cambridge, UK) in a 1:5 ratio and incubated for 30 min to separate leukocytes from erythrocytes. .. Subsequently, the white-coloured layer of leukocytes was transferred to a fresh tube and neutrophils were isolated using the direct human neutrophil isolation kit (Stemcell) according to manufacturer’s instructions. .. Biotinylated antigen was coupled to yellow-green Neutravidin beads (Life Technologies, Thermo Fisher Scientific).

Article Title: Evolution of functional antibodies following acute Epstein-Barr virus infection
Article Snippet: For the isolation of primary human neutrophils from healthy donor blood leucocytes were separated from erythrocytes by mixing the blood in a 5:1 ratio with Hetasep (Stemcell Technologies). .. After an incubation of 15 min at 37°C the leucocytes were washed (500xg, 10 min, 4°C) with MACS buffer (PBS, pH 7.2 + 0.5% BSA + 2mM EDTA, sterile filtered) and purified using the direct human neutrophil isolation kit (Stemcell Technologies). ..

Article Title: A leucine‐rich repeat kinase 2 ( LRRK2 ) pathway biomarker characterization study in patients with Parkinson's disease with and without LRRK2 mutations and healthy controls
Article Snippet: .. For neutrophil isolation, whole blood was collected in a K2EDTA tube and neutrophil isolation was performed within 1 h using a Direct Human Neutrophil Isolation Kit and RoboSep device (StemCell) following the manufacturer's protocol. .. Frequency of neutrophils in whole blood, yield, and purity (CD16+, CD66b+, CD45+) of the negative fraction containing the neutrophils were assessed by flow cytometry using a MACSQuant 10 analyzer (Miltenyi Biotec).

Article Title: A Designed Host Defense Peptide for the Topical Treatment of MRSA-Infected Diabetic Wounds.
Article Snippet: .. Neutrophils were isolated from whole blood via negative enrichment using a direct human neutrophil isolation kit (EasySepTM, STEMCELL Technologies, Cambridge, MA, USA). ..



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STEMCELL Technologies Inc easyseptm direct human neutrophil isolation kit
Easyseptm Direct Human Neutrophil Isolation Kit, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/product/direct+human+neutrophil+isolation+kit/easysep+human+t+cell+isolation+kit/pm40664875-183-24-37
Average 90 stars, based on 1 article reviews
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A) Cartoon illustration of the chemotaxis egg shaped device (see method section for full description). B) The percentage of migration of untreated and iRBC-EVs treated primary neutrophils toward fMLP gradients using the egg-shaped device in A. C) Graph and images showing the uniformity in the size of each patterned C. albicans spot in the swarming assay device . D) Sequential images showing accumulation of untreated neutrophils and iRBC-EVs treated neutrophils at the swarm site at 0, 60, 120, 240, 480 and 600 minutes around zymosan particles (scale bar 25 μm). E) The dynamics of <t>neutrophil</t> swarm size in untreated neutrophils (blue line) and iRBC-EVs treated neutrophils (orange line). Fi) Sequential images showing C. albicans growth in absence of neutrophils at 0mins, 120mins, 240mins, 480mins and 720 mins (scale bar 100 μm). C. albicans expresses a far-red fluorescent protein. The intensity of the red dye is used to assess growth. Transformed black and white images are shown. ii) Sequential images showing the killing of C. albicans (far-red fluorescent protein) and swarm formation (blue, nuclei of neutrophils): untreated primary neutrophils. iii) iRBC-EVs treated primary neutrophils around C. albicans at 0mins, 120mins, 240mins, 480mins and 720 mins (Scale bar 100 μm, n =10 experiments). Gi) The growth dynamics of C. albicans with untreated primary neutrophils (blue line) and iRBC-EVs treated primary neutrophils (orange line). ii) The dynamics of primary neutrophils swarming. Untreated primary neutrophils (blue curve) and iRBC-EVs treated primary neutrophils (orange curve). N=60 swarms across three donors for figures E and G except for the control group. Error bars represent mean +/- standard error for these measurements.
Human Neutrophil Direct Isolation Kit, supplied by STEMCELL Technologies Inc, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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A) Cartoon illustration of the chemotaxis egg shaped device (see method section for full description). B) The percentage of migration of untreated and iRBC-EVs treated primary neutrophils toward fMLP gradients using the egg-shaped device in A. C) Graph and images showing the uniformity in the size of each patterned C. albicans spot in the swarming assay device . D) Sequential images showing accumulation of untreated neutrophils and iRBC-EVs treated neutrophils at the swarm site at 0, 60, 120, 240, 480 and 600 minutes around zymosan particles (scale bar 25 μm). E) The dynamics of neutrophil swarm size in untreated neutrophils (blue line) and iRBC-EVs treated neutrophils (orange line). Fi) Sequential images showing C. albicans growth in absence of neutrophils at 0mins, 120mins, 240mins, 480mins and 720 mins (scale bar 100 μm). C. albicans expresses a far-red fluorescent protein. The intensity of the red dye is used to assess growth. Transformed black and white images are shown. ii) Sequential images showing the killing of C. albicans (far-red fluorescent protein) and swarm formation (blue, nuclei of neutrophils): untreated primary neutrophils. iii) iRBC-EVs treated primary neutrophils around C. albicans at 0mins, 120mins, 240mins, 480mins and 720 mins (Scale bar 100 μm, n =10 experiments). Gi) The growth dynamics of C. albicans with untreated primary neutrophils (blue line) and iRBC-EVs treated primary neutrophils (orange line). ii) The dynamics of primary neutrophils swarming. Untreated primary neutrophils (blue curve) and iRBC-EVs treated primary neutrophils (orange curve). N=60 swarms across three donors for figures E and G except for the control group. Error bars represent mean +/- standard error for these measurements.

Journal: bioRxiv

Article Title: Plasmodium falciparum subverts neutrophil function via host miR-451a loaded extracellular vesicles driving bacterial superinfection susceptibility

doi: 10.1101/2025.06.02.657463

Figure Lengend Snippet: A) Cartoon illustration of the chemotaxis egg shaped device (see method section for full description). B) The percentage of migration of untreated and iRBC-EVs treated primary neutrophils toward fMLP gradients using the egg-shaped device in A. C) Graph and images showing the uniformity in the size of each patterned C. albicans spot in the swarming assay device . D) Sequential images showing accumulation of untreated neutrophils and iRBC-EVs treated neutrophils at the swarm site at 0, 60, 120, 240, 480 and 600 minutes around zymosan particles (scale bar 25 μm). E) The dynamics of neutrophil swarm size in untreated neutrophils (blue line) and iRBC-EVs treated neutrophils (orange line). Fi) Sequential images showing C. albicans growth in absence of neutrophils at 0mins, 120mins, 240mins, 480mins and 720 mins (scale bar 100 μm). C. albicans expresses a far-red fluorescent protein. The intensity of the red dye is used to assess growth. Transformed black and white images are shown. ii) Sequential images showing the killing of C. albicans (far-red fluorescent protein) and swarm formation (blue, nuclei of neutrophils): untreated primary neutrophils. iii) iRBC-EVs treated primary neutrophils around C. albicans at 0mins, 120mins, 240mins, 480mins and 720 mins (Scale bar 100 μm, n =10 experiments). Gi) The growth dynamics of C. albicans with untreated primary neutrophils (blue line) and iRBC-EVs treated primary neutrophils (orange line). ii) The dynamics of primary neutrophils swarming. Untreated primary neutrophils (blue curve) and iRBC-EVs treated primary neutrophils (orange curve). N=60 swarms across three donors for figures E and G except for the control group. Error bars represent mean +/- standard error for these measurements.

Article Snippet: Primary neutrophils were isolated within 1h after draw using the human neutrophil direct isolation kit (STEMcell Technologies) following the manufacturer’s instructions.

Techniques: Chemotaxis Assay, Migration, Transformation Assay, Control

A) Transmission Electron Microscopy image of iRBC-EV. The bar represents a size of 1 μm, B) Size distribution of iRBC EVs as determined by Nanoparticle Tracking Analysis showing the size distribution of iRBC-EV. C) Confocal microscopy of EV uptake by neutrophils. Isolated neutrophils were incubated with 100μg of PKH-67 fluorescently labelled EVs for 1h at 37°C. Neutrophils were stained for actin (Phalloidin, red) and nuclei (Hoechst-blue). D) Representative image from Amnis ImageStream System analysis based on the green fluorescent intensity, we divided into high EV intensity (Yellow colored-top image) and low EV intensity (Orange colored-bottom image) to exclude debris. Representative images from Amnis ImageStream System analysis showing neutrophils (brightfield) interacting with D) fluorescently labeled iRBC-EVs (green, PKH67) (High EV intensity) and D) fluorescently labeled iRBC-EVs (green, PKH67) (low EV intensity). E) Representative image from Amnis ImageStream System analysis showing quantification of low EV intensity internalized by neutrophils. E) Representative image from Amnis ImageStream System analysis showing neutrophils (brightfield) with 0, 1 or 2 internalized fluorescently labeled iRBC-EVs (green, PKH67) (low EV intensity). Representative results from at least three experiments are shown. F) Effect of endocytosis and cytoskeletal reorganization inhibitors on EV uptake. Cells were pretreated for 30 min, with the inhibitors before adding the EVs for 2h. i) Control, ii) Dynasore (80 μM), iii) Colchicine (31 nM), iv) Latrunculin A (5 μM) and iii) Nocodazole (20 μM). One representative experiment of 3 independent experiments is shown. G) effect of inhibitors measured as number of EVs per cell (median; n =3 experiments). Comparisons versus the untreated control were performed using a one-way ANOVA with the Kruskla-Wallis test. Significance levels are indicated as * P <0.05, ** P <0.01 and *** P <0.001.

Journal: bioRxiv

Article Title: Plasmodium falciparum subverts neutrophil function via host miR-451a loaded extracellular vesicles driving bacterial superinfection susceptibility

doi: 10.1101/2025.06.02.657463

Figure Lengend Snippet: A) Transmission Electron Microscopy image of iRBC-EV. The bar represents a size of 1 μm, B) Size distribution of iRBC EVs as determined by Nanoparticle Tracking Analysis showing the size distribution of iRBC-EV. C) Confocal microscopy of EV uptake by neutrophils. Isolated neutrophils were incubated with 100μg of PKH-67 fluorescently labelled EVs for 1h at 37°C. Neutrophils were stained for actin (Phalloidin, red) and nuclei (Hoechst-blue). D) Representative image from Amnis ImageStream System analysis based on the green fluorescent intensity, we divided into high EV intensity (Yellow colored-top image) and low EV intensity (Orange colored-bottom image) to exclude debris. Representative images from Amnis ImageStream System analysis showing neutrophils (brightfield) interacting with D) fluorescently labeled iRBC-EVs (green, PKH67) (High EV intensity) and D) fluorescently labeled iRBC-EVs (green, PKH67) (low EV intensity). E) Representative image from Amnis ImageStream System analysis showing quantification of low EV intensity internalized by neutrophils. E) Representative image from Amnis ImageStream System analysis showing neutrophils (brightfield) with 0, 1 or 2 internalized fluorescently labeled iRBC-EVs (green, PKH67) (low EV intensity). Representative results from at least three experiments are shown. F) Effect of endocytosis and cytoskeletal reorganization inhibitors on EV uptake. Cells were pretreated for 30 min, with the inhibitors before adding the EVs for 2h. i) Control, ii) Dynasore (80 μM), iii) Colchicine (31 nM), iv) Latrunculin A (5 μM) and iii) Nocodazole (20 μM). One representative experiment of 3 independent experiments is shown. G) effect of inhibitors measured as number of EVs per cell (median; n =3 experiments). Comparisons versus the untreated control were performed using a one-way ANOVA with the Kruskla-Wallis test. Significance levels are indicated as * P <0.05, ** P <0.01 and *** P <0.001.

Article Snippet: Primary neutrophils were isolated within 1h after draw using the human neutrophil direct isolation kit (STEMcell Technologies) following the manufacturer’s instructions.

Techniques: Transmission Assay, Electron Microscopy, Confocal Microscopy, Isolation, Incubation, Staining, Labeling, Control

A) ROS-production by unstimulated, PMA and iRBC EVs + PMA treated primary neutrophils stimulated with PMA. One representative experiment of 3 independent experiments performed in triplicates is shown. B) To determine the anti-bacterial activity, isolated neutrophils were pre-exposed to EVs or not and then incubated with S. typhimurium before assessing bacterial load by colony forming unit (CFU) assay. Data are expressed as means ± SEM of 3 independent experiments performed in triplicates. Statistical significance was determined using the Mann-Whitney unpaired test, with *P < 0.05, **P < 0.01, and ***P < 0.001. C) Representative images used for NET quantifications, showing staining for trapped extracellular DNA (green) in response to PMA (100nM), iRBC-EVs(100μg) and PMA/iRBC-EVs in whole blood from healthy donors ( n = 4). Scale bars, 50 μm D ) Quantification of NETosis in whole blood from healthy donors ( n = 4) in response to PMA (100nM), iRBC-EVs(100μg) and PMA/iRBC-EVs. Asterisks indicate significance: *** P < 0.001 by two-way analysis of variance (ANOVA). E) Representative images showing staining for DNA (blue) and histone (green) in response to PMA (100nM), iRBC-EVs (100 μg) and PMA/iRBC-EVs in PMNs isolated from healthy donors ( n = 4) Scale bars, 20 μm.

Journal: bioRxiv

Article Title: Plasmodium falciparum subverts neutrophil function via host miR-451a loaded extracellular vesicles driving bacterial superinfection susceptibility

doi: 10.1101/2025.06.02.657463

Figure Lengend Snippet: A) ROS-production by unstimulated, PMA and iRBC EVs + PMA treated primary neutrophils stimulated with PMA. One representative experiment of 3 independent experiments performed in triplicates is shown. B) To determine the anti-bacterial activity, isolated neutrophils were pre-exposed to EVs or not and then incubated with S. typhimurium before assessing bacterial load by colony forming unit (CFU) assay. Data are expressed as means ± SEM of 3 independent experiments performed in triplicates. Statistical significance was determined using the Mann-Whitney unpaired test, with *P < 0.05, **P < 0.01, and ***P < 0.001. C) Representative images used for NET quantifications, showing staining for trapped extracellular DNA (green) in response to PMA (100nM), iRBC-EVs(100μg) and PMA/iRBC-EVs in whole blood from healthy donors ( n = 4). Scale bars, 50 μm D ) Quantification of NETosis in whole blood from healthy donors ( n = 4) in response to PMA (100nM), iRBC-EVs(100μg) and PMA/iRBC-EVs. Asterisks indicate significance: *** P < 0.001 by two-way analysis of variance (ANOVA). E) Representative images showing staining for DNA (blue) and histone (green) in response to PMA (100nM), iRBC-EVs (100 μg) and PMA/iRBC-EVs in PMNs isolated from healthy donors ( n = 4) Scale bars, 20 μm.

Article Snippet: Primary neutrophils were isolated within 1h after draw using the human neutrophil direct isolation kit (STEMcell Technologies) following the manufacturer’s instructions.

Techniques: Activity Assay, Isolation, Incubation, Colony-forming Unit Assay, MANN-WHITNEY, Staining

A) Experimental setup (n=3 for each stimulation). B) PCA of all samples using the full transcriptome. Shown in parentheses on the axes is the percentage of variance explained by each of the principal components. C) Enrichment analysis of DEGs in neutrophils stimulated with P. falciparum EVs compared to unstimulated neutrophils. Enrichment dot plot of the top Gene Ontology (GO) terms enriched among DEGs. Gene ratio: proportion of DEGs associated with each GO term relative to the total number of genes in that term. Enriched GO terms in y-axis are ordered by adjusted p-value (p.adjust). Dots size: number of DEGs contributing to each term. Dot color: adjusted p-value, with darker colors indicating more statistically significant terms. D) Volcano plot of DEGs in neutrophils stimulated with LPSEV compared to Grouped lpsev condition. All dots correspond to 71 genes with 20% change in expression ((LFC = log2(1.2). Significantly upregulated genes (red dots) and significantly downregulated genes (blues dots). The dashed horizontal line indicates the threshold for statistical significance (−log10(0.05)), and the vertical dashed lines represent the log2 fold change thresholds of 1.2 and −1.2. The labeled genes in blue and the corresponding black dot are genes involve in ferroptosis. E) Box plots display the expression levels of ferroptosis-related genes (HMOX1, KEAP1, NQO1, GPAT4, SLC7A11, and TGFB1) across different conditions (x-axis). The y-axis represents normalized gene expression values.

Journal: bioRxiv

Article Title: Plasmodium falciparum subverts neutrophil function via host miR-451a loaded extracellular vesicles driving bacterial superinfection susceptibility

doi: 10.1101/2025.06.02.657463

Figure Lengend Snippet: A) Experimental setup (n=3 for each stimulation). B) PCA of all samples using the full transcriptome. Shown in parentheses on the axes is the percentage of variance explained by each of the principal components. C) Enrichment analysis of DEGs in neutrophils stimulated with P. falciparum EVs compared to unstimulated neutrophils. Enrichment dot plot of the top Gene Ontology (GO) terms enriched among DEGs. Gene ratio: proportion of DEGs associated with each GO term relative to the total number of genes in that term. Enriched GO terms in y-axis are ordered by adjusted p-value (p.adjust). Dots size: number of DEGs contributing to each term. Dot color: adjusted p-value, with darker colors indicating more statistically significant terms. D) Volcano plot of DEGs in neutrophils stimulated with LPSEV compared to Grouped lpsev condition. All dots correspond to 71 genes with 20% change in expression ((LFC = log2(1.2). Significantly upregulated genes (red dots) and significantly downregulated genes (blues dots). The dashed horizontal line indicates the threshold for statistical significance (−log10(0.05)), and the vertical dashed lines represent the log2 fold change thresholds of 1.2 and −1.2. The labeled genes in blue and the corresponding black dot are genes involve in ferroptosis. E) Box plots display the expression levels of ferroptosis-related genes (HMOX1, KEAP1, NQO1, GPAT4, SLC7A11, and TGFB1) across different conditions (x-axis). The y-axis represents normalized gene expression values.

Article Snippet: Primary neutrophils were isolated within 1h after draw using the human neutrophil direct isolation kit (STEMcell Technologies) following the manufacturer’s instructions.

Techniques: Expressing, Labeling, Gene Expression

A) EV uptake does not induce miRNA expression in neutrophils. Neutrophils, +/- pretreatment with α-amanitin for 30 minutes were incubated with EVs or left untreated. miR451a was quantified by qPCR in neutrophils upon EV incubation. qPCR results from all experiments are normalized by the 2 -Ct method, using RNU6 as a reference and expressed as mean and fold induction over control (mean ± s.e.m.; n =3 experiments), P versus control (Student’s t -test). B) Detection of miRNA by RNA FISH. miRNA transfer to neutrophils by EVs is measured by RNA FISH. miR451a is only detected in neutrophils upon EV treatment, and only when a sequence-specific probe is used for detection (green spots: single-molecule RNA FISH; maximum intensity merges of Z-stacks). RNU6, positive control; scramble, negative control. Blue, nuclei (DAPI). P versus control (Student’s t-test). C) dHL60 expressing miR451 or scramble RNA were incubated with Staphylococcus aureus expressing GFP and phagocytosis was measured by fluorescent microscopy. D) To determine the antibacterial activity of dHL-60 miR451a and dHL-60 scramble , the cells were incubated with S. typhimurium E) S. aureus in a CFU time course experiment. Viable bacteria colonies were counted after incubation (mean ± s.e.m.). N=9 from 3 independent experiments. Comparisons between dHL-60 miR451a and dHL-60 scramble at each time point were performed using a two-way ANOVA with the Sídák’s multiple comparisons test. Significance levels are indicated as * P <0.05, ** P <0.01, *** P <0.001 and **** P <0.0001.

Journal: bioRxiv

Article Title: Plasmodium falciparum subverts neutrophil function via host miR-451a loaded extracellular vesicles driving bacterial superinfection susceptibility

doi: 10.1101/2025.06.02.657463

Figure Lengend Snippet: A) EV uptake does not induce miRNA expression in neutrophils. Neutrophils, +/- pretreatment with α-amanitin for 30 minutes were incubated with EVs or left untreated. miR451a was quantified by qPCR in neutrophils upon EV incubation. qPCR results from all experiments are normalized by the 2 -Ct method, using RNU6 as a reference and expressed as mean and fold induction over control (mean ± s.e.m.; n =3 experiments), P versus control (Student’s t -test). B) Detection of miRNA by RNA FISH. miRNA transfer to neutrophils by EVs is measured by RNA FISH. miR451a is only detected in neutrophils upon EV treatment, and only when a sequence-specific probe is used for detection (green spots: single-molecule RNA FISH; maximum intensity merges of Z-stacks). RNU6, positive control; scramble, negative control. Blue, nuclei (DAPI). P versus control (Student’s t-test). C) dHL60 expressing miR451 or scramble RNA were incubated with Staphylococcus aureus expressing GFP and phagocytosis was measured by fluorescent microscopy. D) To determine the antibacterial activity of dHL-60 miR451a and dHL-60 scramble , the cells were incubated with S. typhimurium E) S. aureus in a CFU time course experiment. Viable bacteria colonies were counted after incubation (mean ± s.e.m.). N=9 from 3 independent experiments. Comparisons between dHL-60 miR451a and dHL-60 scramble at each time point were performed using a two-way ANOVA with the Sídák’s multiple comparisons test. Significance levels are indicated as * P <0.05, ** P <0.01, *** P <0.001 and **** P <0.0001.

Article Snippet: Primary neutrophils were isolated within 1h after draw using the human neutrophil direct isolation kit (STEMcell Technologies) following the manufacturer’s instructions.

Techniques: Expressing, Incubation, Control, Sequencing, Positive Control, Negative Control, Microscopy, Activity Assay, Bacteria