ERG was recorded by using the electrophysiological RETI-animal (Roland Consult, Brandenburg, Germany) system as described by us20 (link),22 (link). Briefly, mice were adapted to darkness for 12h. All of the following procedures were performed under deep red illumination. The animals were anaesthetized intraperitoneally with a mixture of Dormitor (1mg/kg medetomidine hydrochloride; Pfizer, UK) and Ketamine. The mice were then placed on a heated platform to keep their body temperature constant (37±0.5 °C) during the measurements. Mouse pupils were dilated using a single drop of 1% Mydriacyl (Alcon, Ontario, Canada). Flash ERG was measured using a gold wire corneal electrode, a forehead reference electrode, and a ground electrode near the tail. A scotopic ERG was obtained from dark-adapted animals at the following increasing light intensities: 0.01 and 3cd-s/m2. A photopic ERG was recorded following 10-minute light adaptation on the background light intensity of 3cd-s/m2. ERG b-wave was measured from the trough of the a-wave to the peak of the positive wave or, when the a-wave was not present, from baseline to the peak of the first positive wave.
Dormitor
Manufactured by Pfizer
Sourced in United Kingdom, United States
Dormitor is a laboratory equipment product designed for cell culture applications. It provides a controlled environment for the maintenance and growth of cells in a laboratory setting. The core function of Dormitor is to maintain stable temperature, humidity, and atmospheric conditions necessary for optimal cell culture.
Automatically generated - may contain errors
Lab products found in correlation
Medetomidine, by Pfizer (2 mentions)
Mydriacyl, by Alcon (2 mentions)
Stereotactic head frame, by Stoelting (2 mentions)
Ketalar, by Pfizer (2 mentions)
Tropicamide 1 eye drops, by Bausch & Lomb (2 mentions)
Phenylephrine 2.5 eye drops, by Bausch & Lomb (2 mentions)
Ketamine, by Zoetis (2 mentions)
Medetomidine hydrochloride, by Pfizer (2 mentions)
Reti animal, by Roland Consult (1 mentions)
Wistar rat, by Janvier Labs (1 mentions)
Doxycycline hydrochloride, by Merck Group (1 mentions)
Phosphate buffered saline (pbs), by Harvard Bioscience (1 mentions)
Fetal bovine serum (fbs), by Harvard Bioscience (1 mentions)
Glutamine, by Harvard Bioscience (1 mentions)
Hygromycin b, by Carl Roth (1 mentions)
Blasticidin s, by InvivoGen (1 mentions)
Atp disodium salt hydrate, by Merck Group (1 mentions)
Goat antihuman fcγ, by Jackson ImmunoResearch (1 mentions)
Adp sodium salt, by Merck Group (1 mentions)
Erg machine, by Diagnosys (1 mentions)
14 protocols using dormitor
1
Electrophysiological Recording of Mouse ERG
2
Stereotactic Delivery of rAAV2/7 in Rat SN
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All animal experiments were carried out in accordance with the European Communities Council Directive of 24 November 1986 (86/609/EEC) and approved by the Bioethical Committee of the KU Leuven (Belgium). Young adult female Wistar rats (Janvier, France) weighing about 200-250 g were housed under a normal 12 h light/dark cycle with free access to pelleted food and tap water. All surgical procedures were performed using aseptic techniques and ketamine (60 mg/kg ip., Ketalar ® , Pfizer, Belgium) and medetomidine (0.4 mg/kg, Dormitor ® , Pfizer) anaesthesia. Following anaesthesia the rodents were placed in a stereotactic head frame (Stoelting, IL, USA). Injections were performed with a 30-gauge needle and a 10-µl Hamilton syringe. All animals were injected with 3 µl rAAV2/7A53Tsynuclein or rAAV2/7 eGFP as a control (3.0E+11 GC/ml). Stereotactic coordinates used for the SN were anteroposterior (AP) -5.3, lateral (LAT) -2.0, dorsoventral (DV) -7.2, calculated from the dura using bregma as reference. The injection rate was 0.25 µl/min, the needle was left in place for an additional 5 min before being retracted.
3
Recombinant Protein Purification and Characterization
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Standard laboratory chemicals were purchased from Carl Roth. Ham's F‐12 growth medium, fetal bovine serum, PBS, and glutamine were from Biochrom. Blasticidin S was obtained from InvivoGen. Hygromycin B came from Carl Roth. ProCHO4 growth medium was from Lonza. All enzymes for cloning were bought from New England Biolabs. Doxycycline hydrochloride, ADP sodium salt, and ATP disodium salt hydrate were purchased from Sigma‐Aldrich. Part of the recombinant hirudin was a kind gift from Prof. Christian P. Sommerhoff (University of Munich), and another part was purchased from Celgene. Collagen‐related peptide was synthesized at AnaSpec and chemically cross‐linked. DiOC6 (3,3′‐dihexyloxacarbocyanine iodide) was from Life Technologies. midazolam (Dormicum; Roche) was purchased from Roche, and medetomidine (Dormitor; Pfizer) and fentanyl were both from Janssen‐Cilag. Recombinant soluble human CD39 (solCD39) was obtained from R&D Systems. Goat–antihuman Fcγ and goat–antihuman IgG (H+L)‐POD (peroxidase) were purchased from Jackson ImmunoResearch.
4
Splenic Nerve Thermoablation in Animal Model
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All animals received an intraperitoneal injection with anesthetic, a mixture of ketamine (Ketamidor, Richeter Pharma AG, Austria) and medetomidine (Dormitor, 1 mg/kg, Pfizer, USA). Once anesthetized, buprenorphine (Bupaq, 0.05 mg/kg) was also administrated subcutaneously for analgesia. Vaseline was applied onto the eyes to prevent drying and corneal damage. A heat lamp and a surgical blanket kept the body temperature from falling. After an abdominal midline incision, the stomach was retracted upwards, and the spleen was stabilized using retractors. The splenic artery was exposed. A thermal cautery was carefully placed near the splenic artery, for 2–3 s until it dilated, resulting in thermoablation of the splenic nerve [12 ]. The abdominal cavity was bathed with warmed saline 0.9%. Sham animals had the same surgery, except for the thermoablation step. Abdominal muscles and skin were sutured at the end of the surgical procedure. Atipamezole (Antisedan (Orion Corporation, Finland)) was administered subcutaneously to revert the anesthetic state.
5
Splenectomy and Spinal Cord Injury Protocol
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All animals received an intraperitoneal injection with anesthetic, a mixture of ketamine (Ketamidor, Richeter Pharma AG, Austria) and medetomidine (Dormitor, 1 mg/kg, Pfizer, USA). Once anesthetized, buprenorphine (Bupaq, 0.05 mg/kg) was also administrated subcutaneously for analgesia. Vaseline was applied onto the eyes to prevent drying and corneal damage. A heat lamp and a surgical blanket kept the body temperature from falling. After lateral left flank incision, the spleen was exposed and removed with forceps and scissors. The abdominal cavity was bathed with warmed saline 0.9%. Sham animals had the spleen exposed but was not removed. Abdominal muscles and skin were sutured at the end of the surgical procedure. SCI (or laminectomy only) was applied after the splenectomy. Atipamezole (Antisedan (Orion Corporation, Finland)) was administered subcutaneously to revert the anesthetic state.
6
Stereotactic Injection of rAAV2/7 α-Synuclein in Mice
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All surgical procedures were performed using aseptic techniques and mice were treated with ketamine (60 mg/kg, i.p., Ketalar, Pfizer, Puurs) and medetomidine (0.4 mg/kg, Dormitor, Pfizer) anaesthesia. Following anaesthesia, the rodents were placed in a stereotactic head frame (Stoelting, Wood Dale, IL). Injections were performed with a 30-gauge needle and a 10-μl Hamilton syringe (Hamilton, Bonaduz, GR, Switzerland). LRRK2 GS or WT mice were injected with 2 μl rAAV2/7 αSYN vector (CMVenh-synapsin-intron-Hs αSyn WT) at a dose of 3 × 109 genome copies/ml. The right SNpc was targeted using the following stereotactic coordinates: anteroposterior − 3.1 mm, mediolateral − 1.2 mm, and dorsoventral 4.3 mm (counting from the skull) using bregma as reference. The injection rate was 0.25 μl/min, with a 5-min interval following initial needle placement. The needle was left in place for an additional 5 min period after the injection procedure before being slowly retracted.
7
Electroretinogram Recordings in Mice
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ERGs were recorded with an Espion ERG Diagnosys machine (Diagnosys, Littleton, MA) as previously described by us18 (link). In brief, mice were anaesthetized intraperitoneally with a mixture of Dormitor (1 mg/kg medetomidine hydrochloride; Pfizer, UK) and Ketamine after overnight dark adaption. Pupils were dilated with 1% Mydriacyl (Alcon, Ontario, Canada). Flash ERG was measured using a gold wire corneal electrode, a forehead reference electrode, and a ground electrode near the tail.
Scotopic, rod-mediated responses were obtained from dark-adapted animals at the following increasing light intensities: 0.01 and 3 cd-s/m2. Photopic, cone-mediated responses were performed following 10-minute light adaptation on the background light intensity of 30 cd/m2. Recordings were obtained at the light intensity of 3 cd-s/m2. The ERG a-wave amplitudes were measured from the baseline to the negative peak and the b-wave was measured from the trough of the a-wave to the peak of the positive wave or, when the a-wave was not present, from baseline to the peak of the first positive wave.
Scotopic, rod-mediated responses were obtained from dark-adapted animals at the following increasing light intensities: 0.01 and 3 cd-s/m2. Photopic, cone-mediated responses were performed following 10-minute light adaptation on the background light intensity of 30 cd/m2. Recordings were obtained at the light intensity of 3 cd-s/m2. The ERG a-wave amplitudes were measured from the baseline to the negative peak and the b-wave was measured from the trough of the a-wave to the peak of the positive wave or, when the a-wave was not present, from baseline to the peak of the first positive wave.
8
Anesthesia for Rat Studies
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Adult male Dark Agouti rats (200–250 g) (n=50) were maintained in a 12-hour light/12-hour dark cycle, and provided standard food and water ad libitum. Animals were anaesthetised by intraperitoneal administration of ketamine (37.5%) (Ketaset; Fort Dodge Animal Health) and medetomidine (25%) (Dormitor; Pfizer, Exton, PA, USA) at 2 ml/kg, except for IOP measurements, for which animals were anaesthetised by 0.4% isoflurane in oxygen.
9
Mice Models for Ophthalmic Research
10
Mice Models for Ophthalmic Research
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Wild type (WT) C57BL/6 mice were provided by the Biomedical Sciences division, University of Oxford. C3H/HeNHsd-Pde6brd1 (herein referred to as Pde6brd1/rd1) mice were purchased from Harlan Laboratories (Hillcrest, UK). Founder Abca4 knockout mice (129S4/SvJae-Abca4tm1Ght/tm1Ght, herein referred to as Abca4-/-) were generously provided by Gabriel Travis, David Geffen School of Medicine, University of California, Los Angeles, USA.(36 (link)) and bred locally at the University of Oxford.
Animals were housed under a 12 hour light (<100 lux) / dark cycle, with food and water available ad libitum. All procedures were performed under the approval of local and national ethical and legal authorities and in accordance with the Association for Research in Vision and Ophthalmology statements on the care and use of animals in ophthalmic research.
At the time of intraocular injection, mice were between 6 and 8 weeks old. For surgery and in vivo imaging procedures, animals were anesthetized by intraperitoneal injection of 1 mg/kg medetomidine (Dormitor 1 mg/mL, Pfizer, Sandwich, UK) and 60mg/kg ketamine (Ketaset 100mg/mL, Fort Dodge, Southampton, UK) and pupils fully dilated with tropicamide 1% eye drops (Bausch & Lomb, Kingston-Upon-Thames, UK) and, for the imaging procedure, phenylephrine 2.5% eye drops (Bausch & Lomb, Kingston-Upon-Thames, UK).
Animals were housed under a 12 hour light (<100 lux) / dark cycle, with food and water available ad libitum. All procedures were performed under the approval of local and national ethical and legal authorities and in accordance with the Association for Research in Vision and Ophthalmology statements on the care and use of animals in ophthalmic research.
At the time of intraocular injection, mice were between 6 and 8 weeks old. For surgery and in vivo imaging procedures, animals were anesthetized by intraperitoneal injection of 1 mg/kg medetomidine (Dormitor 1 mg/mL, Pfizer, Sandwich, UK) and 60mg/kg ketamine (Ketaset 100mg/mL, Fort Dodge, Southampton, UK) and pupils fully dilated with tropicamide 1% eye drops (Bausch & Lomb, Kingston-Upon-Thames, UK) and, for the imaging procedure, phenylephrine 2.5% eye drops (Bausch & Lomb, Kingston-Upon-Thames, UK).
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