Total protein from cells or tissues was extracted with RIPA (C1053, Applygen). Total protein was quantified by BCA protein assay (23228, Thermo Scientific). All samples were electrophoresed on 10% SDS-PAGE gels before being transferred to a polyvinylidene fluoride membrane (Millipore). Membranes were blocked in 5% skim milk and then incubated with primary antibody overnight at 4 °C. The primary antibodies used in this study were as follows: Ltf (1:250, bs-5810R, Bioss), α-tubulin (B1052, Biodragon, Beijing), β-actin (sc-47778, Santa Cruz). After being treated with the secondary antibodies for 1 h, the protein bands were observed with ECL chemiluminescence solution. The relative intensity of protein signals was normalized to the corresponding loading control intensity and quantified by densitometric analysis with ImageJ software (https://imagej.nih.gov/ij/ ).
Bs 5810r
Manufactured by Bioss Antibodies
Sourced in China
The Bs-5810R is a laboratory centrifuge designed for general-purpose applications. It features a maximum speed of 5,810 rpm and a maximum relative centrifugal force (RCF) of 5,550 x g. The centrifuge accommodates a variety of sample sizes and can be used for tasks such as separating liquids, concentrating samples, and preparing samples for analysis.
Automatically generated - may contain errors
Lab products found in correlation
Goat anti rabbit igg hrp, by Thermo Fisher Scientific (2 mentions)
Goat anti chicken igg hrp, by Thermo Fisher Scientific (2 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
β actin sc 47778, by Santa Cruz Biotechnology (1 mentions)
Bca protein assay, by Thermo Fisher Scientific (1 mentions)
C1053, by Applygen (1 mentions)
Ab2302, by Merck Group (1 mentions)
Edas 290 system, by Kodak (1 mentions)
A11012, by Thermo Fisher Scientific (1 mentions)
Eclipse ti u, by Nikon (1 mentions)
A11001, by Thermo Fisher Scientific (1 mentions)
Tcs sp8 sted, by Leica (1 mentions)
A11005, by Thermo Fisher Scientific (1 mentions)
A11034, by Thermo Fisher Scientific (1 mentions)
Ab45977, by Abcam (1 mentions)
Ab243066, by Abcam (1 mentions)
Bs 0295g af555, by Bioss Antibodies (1 mentions)
5 protocols using bs 5810r
1
Western Blot Analysis of Ltf Protein
2
Quantification of LTF and GAPDH in Striatum
3
Quantitative Western Blot Analysis
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Protein levels were determined using Western blotting, as previously described6 (link). Rabbit anti-lactoferrin (bs-5810R, Bioss), mouse anti-STAT3 (9139 Cell Signaling), rabbit anti-p-STAT3 (9131S, Cell Signaling), or chicken anti-GAPDH (Millipore, AB2302) immunopositive bands were visualized using goat anti-rabbit IgG-HRP (invitrogen) or goat anti-chicken IgG-HRP (Invitrogen) and ECL (Pierce, Rockford, IL) (see Supplementary Table 2 for more information). Semi-quantification of luminescence signal intensity on X-ray film was determined by the analyses of optical density, using a Computer-Assisted Kodak Analysis (EDAS) 290 system. Western blots of Fig. 2g (STAT3) and Fig. 5g (LTF) are included as Supplementary Figs. 2 and 3 , respectively.
4
Immunofluorescence Analysis of Neuroinflammation
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At 1- and 3-days following ICH, the immunofluorescence was performed as previously prescribed [6 ]. Briefly, the frozen coronal slices were put into antigen repair solution and permeabilized with 1% Triton X-100 for 15min. Brain sections were blocked in 5% goat serum and incubated with anti-rabbit MPO (1:500, ab45977, Abcam), anti-rabbit NeuN (1:500, CST), anti-MDA [11E3] (1:200, ab243066, Abcam), anti-rabbit Iba1 (019–19741, Wako), anti-mouse Ly6G (551459, BD Biosciences), anti-mouse CD11b-FITC (Clone:M1/70, 1:200, 4306305, eBioscience), anti-rabbit Ltf (1:200, bs-5810R, Bioss) primary antibody at 4 °C overnight. Slices then incubated with goat anti-rabbit 594 (A11012, Invitrogen), goat anti-rabbit 488 (A11034, Invitrogen), goat anti-mouse 594 (A11005, Invitrogen), goat anti-mouse 488 (A11001, Invitrogen). Nuclei were visualized via a mounting medium containing DAPI (ZG1202, Vectarshield). Images were analyzed with a confocal microscope (TCS SP8 STED, Leica) and fluorescence microscope (ECLIPSE Ti–U, NIKON). Mean fluorescence intensity (MFI) was analyzed with ImageJ software.
5
Immunocytochemical Detection of Lactoferrin in Sperm
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Smears of the final suspension of sperm in diluent were air-dried on microscope slides at room temperature and fixed in methanol, after which the slides were washed three times in PBS for 10 min and blocked for 30 min at room temperature in 5% BSA. Sperm were incubated overnight at 4°C with anti-LTF (BS-5810R; Bioss, China) (1:400). For the negative control, PBS was used instead of primary antibody. Next, sperm were incubated with Alexa Fluor 555-conjugated goat anti-rabbit IgG H&L (1:400) (BS-0295G-AF555; Bioss, China) for 1 h at 37°C. Finally, sperm were incubated in the dark at 37°C for 10 min for staining with DAPI (4′,6-diamidino-2-phenylindole). Images were obtained using a confocal laser microscope (LSM; Zeiss, Oberkochen, Germany); red fluorescence on the images indicated positivity.
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