Journal: Methods and Protocols

Article Title: Sequential Isolation of Microglia and Astrocytes from Young and Aged Adult Mouse Brains for Downstream Transcriptomic Analysis

doi: 10.3390/mps5050077

Figure Lengend Snippet: Detailed information on required materials.

Article Snippet: CD11b (Microglia) MicroBeads, mouse/human (1 mL) Anti-ACSA-2 MicroBead Kit mouse (2 × 1 mL) FcR blocking reagent (1 mL) Anti-ACSA-2 MicroBeads (1 mL) , Miltenyi Biotec B.V. & Co. KG , 130-093-634 , Bergisch Gladbach, DE.

Techniques: Saline, Blocking Assay

Journal: Cell Insight

Article Title: Cannabinoid receptor 2 signal promotes type 2 immunity in the lung

doi: 10.1016/j.cellin.2023.100124

Figure Lengend Snippet: Genetic deletion of CB2 does not affect the normal development or immune homeostasis of the lung. (A to C) Normal lung development with the CB2 deletion. (A) Gross appearance of the lung tissues (left lobe) of adult CB2 −/− and control CB2 +/+ littermates. (B) Tissue weights (left lobe) were measured. n = 6, mean ± SEM, n.s., not significant (Student's t -test). (C) Representative images of the lung tissue sections of CB2 +/+ and CB2 −/− mice assessed by hematoxylin and eosin (H&E) staining. (D) The CB2 deletion does not affect tissue levels of the endocannabinoid 2-AG. 2-AG tissue levels in the lungs of CB2 +/+ and CB2 −/− mice were compared by the ELISA analysis. n = 6, mean ± SEM, n.s., not significant (Student's t -test). (E and F) Immune cell populations are not affected by the CB2 deletion. (E) ILC2s and other immune cell types in the lungs of CB2 +/+ and CB2 −/− mice were examined by the FACS analysis. n = 5, mean ± SEM, n.s., not significant (Student's t -test). (F) Representative FACS gating of eosinophils (CD45 + CD11b + CD11c − Siglec-F + ) and alveolar macrophages (CD45 + CD11b + CD11c + Siglec-F + ) in the lung.

Article Snippet: 2-AG levels in the supernatants were measured by the Mouse 2-AG Elisa Kit (Krishgen Biosystems).

Techniques: Control, Staining, Enzyme-linked Immunosorbent Assay

Journal: Cell Insight

Article Title: Cannabinoid receptor 2 signal promotes type 2 immunity in the lung

doi: 10.1016/j.cellin.2023.100124

Figure Lengend Snippet: Genetic deletion of CB2 mitigates the IL-33-elicited type 2 immunity in the lung. (A and B) Wild-type mice were intranasally treated with IL-33 or control saline. (A) ILC2s were FACS-sorted from the saline-treated or IL-33-treated lung tissues, and CB2 mRNA levels in ILC2s were compared by the qPCR analysis. n = 3, mean ± SEM, n.s., not significant (Student's t -test). (B) 2-AG levels in the saline-treated or IL-33-treated lung tissues were quantified by the ELISA analysis. n ​= ​4, mean ​± ​SEM, ∗ p < 0.05 (Student's t -test). (C to K) CB2 −/− and control CB2 +/+ littermates were subjected to intranasal IL-33 treatment. (C) ILC2s in the IL-33-treated lungs of CB2 +/+ and CB2 −/− mice were examined by the FACS analysis. n = 7, mean ± SEM, n.s., not significant (Student's t -test). (D) mRNA levels of IL-4 , IL-5 , and IL-13 in the lung tissues of CB2 +/+ and CB2 −/− mice were determined by the qPCR analysis and normalized to the saline-treated CB2 +/+ condition. n ​= ​5 for saline-treated condition and n ​= ​15 for IL-33-treated condition, mean ​± ​SEM, ∗ p < 0.05 (two-way ANOVA test). (E) Protein levels of IL-5 and IL-13 in the IL-33-treated lungs of CB2 +/+ and CB2 −/− mice were quantified by the ELISA analysis. n ​= ​3, mean ​± ​SEM, ∗ p < 0.05 (Student's t -test). (F and G) IL-33-elicited immune response in the lung was visualized by advanced 3D imaging. The intact, unsectioned lungs (left lobe) of CB2 +/+ and CB2 −/− mice were processed for the whole-tissue anti-Siglec-F immunolabeling. (F) Representative 3D-projection images at 1.26 × magnification of the lightsheet imaging were shown. (G) The density of Siglec-F + immune cells was quantified. n ​= ​5, mean ​± ​SEM, ∗ p < 0.05 (two-way ANOVA test). (H and I) Eosinophil recruitment in the lungs of CB2 +/+ and CB2 −/− mice was assessed by the FACS analysis. (H) Representative FACS gating of eosinophils (CD45 + CD11b + CD11c − Siglec-F + ) in the IL-33-treated condition. (I) The percentage of eosinophils in total immune cells was quantified. n ​= ​5 for saline-treated condition and n ​= ​12 for IL-33-treated condition, mean ​± ​SEM, ∗ p < 0.05 (two-way ANOVA test). (J and K) The lung tissues of CB2 +/+ and CB2 −/− mice were assessed by hematoxylin and eosin (H&E) staining. (J) Representative images were shown. (K) Histologic scores were determined. n ​= ​5 for saline-treated condition and n ​= ​7 for IL-33-treated condition, mean ​± ​SEM, ∗ p < 0.05 (two-way ANOVA test).

Article Snippet: 2-AG levels in the supernatants were measured by the Mouse 2-AG Elisa Kit (Krishgen Biosystems).

Techniques: Control, Saline, Enzyme-linked Immunosorbent Assay, Imaging, Immunolabeling, Staining