Approaching 72 hours after GMH, rats were given intraperitoneal injections of 2% Evans blue in normal saline (4 ml/kg of body weight) 3 hours before euthanasia, as described previously (Manaenko et al., 2011 (link)). Afterwards, under deep isoflurane anesthesia, rats were transcardially perfused with ice cold PBS. The brain tissue was removed, frozen in liquid nitrogen, and stored at −80 °C until analysis. Brains were homogenized in 1100 µl of PBS, sonicated, centrifuged (30 minutes, 15,000 rcf, 4 °C), and supernatants collected. Hemoglobin assay was performed by adding Drabkin’s reagent (0.8 ml, Sigma-Aldrich) to 0.2 ml supernatant aliquots, which were allowed to stand for 15 minutes at room temperature. Sample absorbance was measured via spectrophotometry (Thermo Spectronic Genesys 10 UV, Thermo Fisher Scientific Inc., Waltham, MA) at 540 nm, and quantified using a standard curve. Results are presented as µl of blood. Additionally, Evans blue assays were performed by adding an equal volume of 50% trichloroacetic acid to supernatant aliquots, after which they were incubated overnight at 4 °C and then centrifuged (30 minutes, 15,000 rcf, 4 °C). Evans blue content was then measured by spectrophotometry at 610 nm and quantified according to a standard curve. The results are presented as (µg of Evans blue dye)/(g of tissue).
Thermo spectronic genesys 10 uv
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Thermo Spectronic Genesys 10 UV is a compact, single-beam UV-Visible spectrophotometer designed for routine laboratory analysis. It is capable of measuring the absorbance of samples in the wavelength range of 190 to 1100 nanometers.
Automatically generated - may contain errors
Lab products found in correlation
7 protocols using thermo spectronic genesys 10 uv
1
Quantitative Analysis of Blood-Brain Barrier Permeability
2
Quantifying BBB Permeability via EB Extravasation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BBB permeability was evaluated by EB extravasation using spectrophotometry as previously described [20 ]. A 2% solution of EB in normal saline (5 ml/kg of body weight) was injected into the right femoral vein at 24 h after SAH. The stain was allowed to circulate for an additional 60 min before sacrifice. Then the rats were transcardially perfused with 120 ml of ice-cold PBS in deep anesthesia. The collected brain samples were subsequently removed and divided into four parts: right hemisphere, left hemispheres, cerebellar, and brain stem and stored in a − 80 °C freezer. After homogenization of each sample with PBS, the solution was centrifuged for 30 min at 14,000 r/min in 4 °C. The supernatant was collected, mixed with equal amount of trichloroacetic acid, and incubated at room temperature for 1 h. Then, the sample was centrifuged for 30 min at 15,000 r/min in 4 °C to separate the supernatant for measurements. EB stain results were measured by a spectrophotometer (Thermo Spectronic Genesys 10 UV, Thermo Fischer Scientific Inc., Waltham, MA, USA) at 610 nm and quantified with a standard curve. The results are presented as fold increase compared to sham group.
3
Evans Blue Dye Quantification in Rat Brains
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Rats in each group were injected with a 2% Evans blue dye solution in saline (4 ml/kg). The dye was allowed to circulate for 2 hours. Cardiac perfusion with phosphate buffered saline (PBS) was performed under deep anesthesia. The brains were then removed, weighed, and homogenized in N, N-dimethylformamide (DMF). The homogenate was maintained at room temperature in the dark for 72 hours and centrifuged at 10,000 × g for 25 minutes. Absorbance of the supernatant was measured using a spectrophotometer (Thermo Spectronic Genesys 10 UV, Thermo Fischer Scientific Inc., Waltham, MA, USA) at 610 nm [13 (link)]. The amount of Evans blue dye was calculated using a standard curve and is expressed as microgram/gram brain tissue.
4
Measuring Blood-Brain Barrier Permeability
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The blood-brain barrier (BBB) permeability was measured by the extravasation of Evans Blue dye on days 1 and 3 post-ICH [22 (link)]. In brief, a 2% solution of Evans Blue in sterile saline (4 ml/kg of body weight, Servicebio, China) was injected into the tail vein 3 h before mice were sacrificed. Then, mice were transcardially perfused with 40 ml of 0.9% cold saline. Afterward, brain tissues were quickly removed, weighed, and homogenized in 1100 μl PBS. After sufficient grounding, the samples were centrifuged at 6000×g for 30 min. The supernatant was collected and mixed with an equal amount of 50% trichloroacetic acid (Sigma-Aldrich, St Louis, MO). Samples were incubated overnight at 4 °C and centrifuged for 30 min (6000×g, 4 °C). Evans Blue dye were measured by a spectrophotometer (Thermo Spectronic Genesys 10 UV, Thermo Fischer Scientific Inc., Waltham, MA, USA) at 610 nm and quantified from a standard curve. The results are presented as (microgram of Evans Blue dye)/(gram of tissue).
5
Quantifying Blood-Brain Barrier Permeability
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Evans Blue (4 mL/kg, 2% solution; Sigma-Aldrich, St. Louis, MO, USA) in saline was injected via the tail vein (6 mice in each group). The dye was allowed to circulate for 2 hours. Mice were then treated with 50 mL ice-cold phosphate buffered saline (PBS) by transcardiac perfusion. The brain was separated into right and left hemispheres and stored at –80°C. Samples were homogenized in 1.1 mL PBS, sonicated, and centrifuged for 30 minutes at 15,000 r/min, 4°C. Supernatants were collected in aliquots, and trichloroacetic acid (50%) added to 500 μL aliquots. The mixture was incubated at 4°C overnight, and then centrifuged for 30 minutes at 15,000 r/min, 4°C. Evans Blue concentration was determined at 620 nm using a spectrophotometer (Thermo Spectronic Genesys 10 UV, Thermo Fisher Scientific Inc., Waltham, MA, USA). Data were quantified from a standard curve. The results are presented as: Evans Blue stain (μg)/tissue (g).
6
Spectrophotometric Brain Haemoglobin Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A modified spectrophotometric haemoglobin assay was used according to the previous study.33 (link) Briefly, the brain was homogenised with 3 mL PBS and centrifuged at 13,000 rpm for 30 min. The supernatant was immediately transferred to a clean tube with a 4:1 ratio of Drabkin's reagent (Sigma-Aldrich, St. Louis, MO, USA) and incubated for 15 min. absorbence of sample was analysed on a standard curve by spectrophotometric measurements at 540 nm wavelength (Thermo Spectronic Genesys 10 UV, Thermo Fischer Scientific).
7
Quantifying Blood-Brain Barrier Permeability
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
A 2% solution of Evans blue in normal saline (4 ml/kg of body weight) was injected i.p. at 24 h after SAH. The stain was allowed to circulate for an additional 24 h before sacrifice. Then, animals were transcardially perfused with 120 ml of ice-cold PBS in deep anesthesia. The collected brain samples were divided into four parts, namely, right hemisphere, left hemisphere, cerebellum, and brain stem, and were stored in a freezer at −80°C. After homogenization of each sample with PBS, the solution was centrifuged for 30 min at 14,000 r/min in 4°C. The supernatant was collected, mixed with an equal amount of trichloroacetic acid, and incubated at room temperature for 1 h. Then, the sample was centrifuged for 30 min at 15,000 r/min in 4°C to separate the supernatant for measurements. Evans blue stain results were measured using a spectrophotometer (Thermo Spectronic Genesys 10 UV, Thermo Fischer Scientific Inc., Waltham, MA, United States) at 610 nm and quantified with a standard curve (Manaenko et al., 2011 (link)). The results are presented as a fold increase compared with the sham group.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!