Clinical formulation of 6mg/ml Paclitaxel Solution for Infusion (Actavis Ltd) was diluted with 0.9% sterile saline (Fresenius Kabi, UK) to obtain a 2mg/ml solution. Control animals received an equivalent amount of vehicle solution. In order to replicate the vehicle solution of the clinical formulation, a 1:1 solution of cremophor EL (Sigma, UK) and ethanol, plus 2mg/ml sodium citrate (Sigma, UK), was used as a vehicle solution. For vehicle administration, the stock vehicle solution was diluted with 0.9% sterile saline, at a ratio of 1:2. 2mg/kg paclitaxel or an equal volume of vehicle solution were administered intraperitoneally on four alternate days (0, 2, 4, 6) [18 (link)].
Sterile saline
Manufactured by Fresenius
Sourced in United Kingdom, Germany
Sterile saline is a salt solution that is typically used for medical and laboratory purposes. It is a clear liquid that is free of particles and is designed to be sterile and pyrogen-free. Sterile saline is commonly used to dilute or reconstitute medications, flush intravenous lines, and clean wounds or other medical equipment.
Automatically generated - may contain errors
Lab products found in correlation
Cremophor el, by Merck Group (4 mentions)
Sodium citrate, by Merck Group (3 mentions)
D mannitol, by Merck Group (2 mentions)
Lipopolysaccharides (lps), by Merck Group (1 mentions)
Sterile saline, by Merck Group (1 mentions)
Mz125, by Leica (1 mentions)
Urethane, by Merck Group (1 mentions)
Forane, by Abbott (1 mentions)
13 protocols using sterile saline
1
Paclitaxel Infusion Solution Preparation
2
Acute LPS Exposure in Rats
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Rats were simultaneously exposed in separate chambers to aerosolised LPS (Sigma-Aldrich, USA) 1mg/ml or sterile saline (Fresenius Kabi, UK) for 30 minutes. At 3 and 24hrs thereafter, rats were euthanised by anaesthetic overdose of intra-peritoneally (i.p) injected Pentobarbitone. Naïve rats used in this study were euthanised by overdose of CO2 inhalation.
3
Bortezomib-Induced Neuropathic Pain Model
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Clinical grade bortezomib (Velcade®) 3.5mg/vial (Millennium Pharmaceuticals, Inc) was diluted with 0.9% sterile saline (Fresenius Kabi, UK) to prepare a 0.2mg/ml bortezomib solution. For vehicle administration, a solution of 0.2mg/ml D-mannitol (Sigma, UK) in 0.9% sterile saline was used to replicate Velcade® vehicle solution. Animals received 0.2mg/kg bortezomib, or an equivalent volume of vehicle solution, intraperitoneally on day 0, 3, 7 and 10 [16 (link)].
Three key time-points were investigated in these models of CIPN. Day 4 (bortezomib) or day 7 (paclitaxel and oxaliplatin), prior to the onset of pain-like behaviour; peak pain, when animals reached the peak of mechanical hypersensitivity (paclitaxel: day 28–31; oxaliplatin: day 27–29; bortezomib: day 23–25); and resolution of chemotherapy-induced pain behaviour, when animals returned to their individual baseline response on two consecutive occasions (paclitaxel day 174; oxaliplatin day 147–148; bortezomib day 116–124). The peak pain and resolution of pain time-points within a cohort occurred on more than one day, as they differed between rats and were dependent on individual rat behaviour compared to its baseline. Raw behavioural data for all animals is shown inS1 Raw data .
Three key time-points were investigated in these models of CIPN. Day 4 (bortezomib) or day 7 (paclitaxel and oxaliplatin), prior to the onset of pain-like behaviour; peak pain, when animals reached the peak of mechanical hypersensitivity (paclitaxel: day 28–31; oxaliplatin: day 27–29; bortezomib: day 23–25); and resolution of chemotherapy-induced pain behaviour, when animals returned to their individual baseline response on two consecutive occasions (paclitaxel day 174; oxaliplatin day 147–148; bortezomib day 116–124). The peak pain and resolution of pain time-points within a cohort occurred on more than one day, as they differed between rats and were dependent on individual rat behaviour compared to its baseline. Raw behavioural data for all animals is shown in
4
Bortezomib-Induced Peripheral Neuropathy Model
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Velcade®, clinical grade bortezomib (Millennium Pharmaceuticals Inc., Takeda Pharmaceutical Company Limited), was used to induce bortezomib‐induced peripheral neuropathy. Powdered Velcade® was reconstituted with 0.9% sterile saline (Fresenius Kabi, UK) to achieve a solution with a final concentration of 0.2 mg·mL−1. A vehicle solution of 0.2 mg·mL−1 D‐mannitol (Sigma, UK), in 0.9% sterile saline, was used to replicate clinical Velcade®. Animals received 0.1 or 0.2 mg·kg−1 bortezomib i.p. or an equal volume of vehicle solution on days 0, 3, 7 and 10. Dosage and administration schedule was based on clinical use of Velcade® [1.3 mg·m−2 (Conversion factor: Nair and Jacob, 2016 ) on days 0, 3, 7 and 10 (Moreau et al.,2012 )]. Velcade® has been shown to induce dose‐dependent proteasome inhibition, with a single dose of 1.38 mg·m−2 yielding 74 ± 2% inhibition, which returned towards baseline after 72 h (More detail: Orlowski et al.,2002 ).
5
Vaginal Cytology Monitoring in Rats
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Vaginal secretion was collected from female rats every morning between 07:00 and 08:00 h for 8–14 consecutive days using a slightly modified protocol described elsewhere (McLean et al., 2012 ). In short: Vaginal lavage was performed by holding the rat in an upright position and placing a filtered pipette tip filled with 100 μl sterile saline (Fresenius Kabi, Bad Homburg, Germany) at the opening of the vaginal canal. 50 μl of the saline was gently released and withdrawn. This procedure was repeated 4–5 times using the same pipette tip. The vaginal sample was placed on a microscope slide and examined under a brightfield microscope (Leica MZ125, Leica Biosystems, Germany). The stage of estrous cycle was determined by the ratio of cells present at time of sample collection. While the proestrous and estrous phases (Pro/Est) are characterized by a high proportion of nucleated epithelial cells (proestrus) and cornified epithelial cells (estrous), metestrus and diestrus (Met/Die) are characterized by a large proportion of leukocytes.
6
Standardized Paclitaxel Administration Protocol
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The clinical formulation of 6 mg/mL Paclitaxel Solution for Infusion (CP Pharmaceuticals Ltd, UK or Actavis Ltd, UK) was diluted with 0.9% sterile saline (Fresenius Kabi, United Kingdom) to achieve a 2 mg/mL solution. To replicate the clinical formulation of paclitaxel, a vehicle stock solution was made consisting of equal parts of cremophor EL (Sigma, Dorset, United Kingdom) and ethanol. In experiments using paclitaxel solution manufactured by Actavis Ltd, the vehicle stock solution also contained 2 mg/mL sodium citrate as per this formulation. For vehicle administration; 1 part vehicle stock solution was diluted with 2 parts 0.9% sterile saline. Rats received intraperitoneal 2 mg/kg paclitaxel or equivalent volume of vehicle solution on 4 alternate days (0, 2, 4, and 6). Animals were dosed according to their weight, ie, 250 g rat received 0.25 mL injection. Injections were performed in the morning, and rats were immediately returned to their home cages afterwards.
7
Paclitaxel Injection Formulation and Administration
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Clinically formulated paclitaxel solution for infusion (6 mg/ml; CP Pharmaceuticals UK or Actavis UK) was diluted with 0.9% sterile saline (Fresenius Kabi, UK) to achieve a 2 mg/ml solution for injection. To replicate the clinical formulation, a vehicle stock solution was made using 1 : 1 solution of Cremophor EL (Sigma, UK) and ethanol. When using paclitaxel from Actavis, the vehicle stock was supplemented with 2 mg/mL sodium citrate (Sigma, UK). Prior to administration, one part of vehicle stock solution was diluted with two parts of 0.9% sterile saline. The rats were dosed with 2 mg/kg paclitaxel or equivalent volume of vehicle solution intraperitoneally on four alternate days (0, 2, 4, and 6). Animals were dosed according to their weight (1 ml/kg) and were immediately returned to their home cages. All paclitaxel/vehicle injections were administered in the early afternoon 1–3 pm. Both paclitaxel and vehicle solutions are clear solutions with the same viscosity, which enables blinding procedures (see below).
8
Paclitaxel-Induced Neuropathic Pain in Rats
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After habituation and baseline mechanical sensitivity testing, rats received intraperitoneal (i.p.) injections of 2 mg/kg paclitaxel on 4 alternate days (days 0, 2, 4, and 6) as previously described.8-11 Paclitaxel concentration for solution for infusion (6 mg/mL; CP Pharmaceuticals Ltd/Actavis, Devon, UK) was diluted with .9% sterile saline (Fresenius Kabi, Manor Park, UK) and injected at 2 mg/mL/kg body weight. Injections were performed in the morning, and rats were immediately returned to their home cages afterward.
9
Impact of Urethane on Neural Activity in P8 Rats
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We recorded from P8 rats before and after injection of urethane (n = 7) or saline (n = 6). We recorded baseline video and neurophysiological data for 30 min while pups cycled between sleep and wake, followed by 50 stimulations of the right forelimb as in Experiment 1. We then infused urethane (1.0 mg/g b.w.; Sigma-Aldrich, St. Louis, MO) or an equivalent volume of sterile saline (Fresenius Kabi, Bad Homburg, Germany) through the implanted cannula. This procedure minimized disruption of the pup and allowed for uninterrupted recording of data. Also, subcutaneous infusion of urethane produces a comparable level of surgical anesthesia as intraperitoneal injection and reduces the likelihood of organ puncture (Maggi and Meli, 1986 (link); Field and Lang, 1988 (link); Matsuura and Downie, 2000 (link)). We waited at least 10 min for the drug to take effect, after which data were again recorded for 30 min followed by 50 stimulations of the right forelimb. One pup was excluded from analysis due to the complete loss of neural activity after urethane administration.
10
Intranasal Exposure to Soy Hull Extract
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Six experimental groups were created. Soy hull extract was resuspended in sterile saline (0.9% NaCl, Fresenius Kabi, Barcelona, Spain) at two different concentrations of 3 and 5 mg protein/ml. Mice were exposed during five consecutive days over three weeks, under light anesthesia with isoflurane (Forane, Abbott Laboratories, Madrid, Spain), to either 20μl of saline, 20μl of SHE at 3 mg protein/ml concentration (Soy3), 20μl of SHE at 5 mg protein/ml concentration (Soy5), or a combination of one of these three solutions with 150 μg of DEP depending on the experimental group. In the groups receiving DEP, the supplement was administered three days a week for three weeks. A chart of the intranasal instillations of the six groups is shown in Fig 1 . Each group comprised eight mice.
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