Journal: Histochemistry and cell biology

Article Title: Choosing the right antibody for Resistin-Like Molecule (RELM/FIZZ) family members

doi: 10.1007/s00418-012-1042-0

Figure Lengend Snippet: Specificity of hRELMβ antibodies in Western blot and immunofluorescence assays. In Western blot assays, hRELMβ antibodies from Acris (A), Adipogen, and Chemicon (B) were each highly specific for hRELMβ when tested against all RELM isoforms, including mouse. (C) Only the goat anti-hRELMβ antibody from Acris exhibited binding to hRELMβ in the immunofluorescence assay. Mouse anti-FLAG antibody from Sigma was used to indicate the inductive expression of fusion protein in HEK 293 cells. Cy2-conjugated donkey anti-goat IgG was used as secondary antibody (C, green)

Article Snippet: hRELMβ , Acris Antibodies Inc. , Peptide (28–41) DSVMDKKIKDVLNS , Goat , X , X.

Techniques: Western Blot, Immunofluorescence, Binding Assay, Expressing

Journal: American journal of physiology. Lung cellular and molecular physiology

Article Title: Hypoxia-induced mitogenic factor (HIMF/FIZZ1/RELMalpha) induces the vascular and hemodynamic changes of pulmonary hypertension.

doi: 10.1152/ajplung.90526.2008

Figure Lengend Snippet: Fig. 1. In vivo knockdown of hypoxia-in- duced mitogenic factor (HIMF) in the chronic hypoxia model of pulmonary hypertension. A: real-time PCR for green fluorescent protein (GFP) from RNA isolated from the lung, heart, liver, and spleen 72 h after intratracheal instillation of Ad-HIMF-shRNA. B: equally divided left lungs from uninfected rats or rats infected with Ad-Neg-shRNA 4 days earlier were homogenized and evaluated for GFP ex- pression. The homogenates were resolved by 4–20% SDS-PAGE and transferred to nitro- cellulose. The blots were probed with rabbit polyclonal anti-GFP antibodies and developed with ECL. To confirm equal protein loading and transfer, the blots were stripped and re- probed with monoclonal anti- -actin antibod- ies. IB, immunoblot; IB*, immunoblot after stripping. C: visualization of GFP in control rat lungs or lungs infected with Ad-Neg- shRNA. DIC, differential interference contrast. D: lung homogenates from animals exposed to 4 days of normoxia (20.8% O2) or hypoxia (10.0% O2) were evaluated for HIMF expression. The homogenates were resolved by 4–20% SDS- PAGE and transferred to nitrocellulose. The blots were probed with polyclonal anti-HIMF antibodies and developed by ECL. To confirm introduction of the viral vectors, the blot was stripped and reprobed with polyclonal anti-GFP antibodies. To confirm equal protein loading, the blots were stripped and reprobed with monoclo- nal anti- -actin antibodies. IB**, immunoblot after second stripping.

Article Snippet: Nonspecific protein binding was blocked by treatment of the slides with normal rabbit serum for 30 min. After the final blocking step, the sections were treated with polyclonal goat anti-mouse RELM (1: 200; R&D Systems, Minneapolis, MN) or antibody diluent alone for 120 min at RT.

Techniques: In Vivo, Knockdown, Real-time Polymerase Chain Reaction, Isolation, shRNA, Infection, SDS Page, Western Blot, Stripping Membranes, Control, Expressing

Journal: American journal of physiology. Lung cellular and molecular physiology

Article Title: Hypoxia-induced mitogenic factor (HIMF/FIZZ1/RELMalpha) induces the vascular and hemodynamic changes of pulmonary hypertension.

doi: 10.1152/ajplung.90526.2008

Figure Lengend Snippet: Fig. 5. Effectiveness of pulmonary HIMF gene transfer. A: 2 wk after rats were intratra- cheally instilled with AAV-HIMF, the lungs were homogenized, resolved by 4–20% SDS- PAGE, and transferred to nitrocellulose. The blots were probed with rabbit anti-HIMF anti- bodies and developed by ECL. To confirm equal loading and transfer, the blot was stripped and reprobed with anti- -actin monoclonal an- tibodies. B: laser densitometry was used to quantify HIMF levels in the lung samples. Data were normalized to -actin expression and ex- pressed as relative intensity (means SE). The number of lungs studied is indicated within each bar. *P 0.05 compared with the control lung. C–E: paraffin-embedded lung sections from AAV-null-treated (C) and AAV-HIMF- treated (D and E) mice were rehydrated and stained with goat anti-mouse HIMF polyclonal antibodies. Scale bar 50 m.

Article Snippet: Nonspecific protein binding was blocked by treatment of the slides with normal rabbit serum for 30 min. After the final blocking step, the sections were treated with polyclonal goat anti-mouse RELM (1: 200; R&D Systems, Minneapolis, MN) or antibody diluent alone for 120 min at RT.

Techniques: SDS Page, Expressing, Control, Staining