Behavioral Tests:
Irwin Test
The classical or modified Irwin...
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- A standard design includes 7 days of acclimatization period followed by test substance dosing and post-dosing periods with observations and testing. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 10-15 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
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Open Field Test
The open-field test is a...
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- A standard design includes 7 days of acclimatization period followed by test substance dosing and post-dosing periods with observations, and the day of testing. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 10-20 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
Wire Hang Test
The Wire Hang test was...
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- A standard design includes 7 days of acclimatization period followed by test substance dosing and post-dosing periods with observations and testing. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 10-15 mice/rats per group, at least two repeated testing procedures (before, and after treatment). Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
Beam Walking
The Beam walking test...
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A standard design includes 7 days of acclimatization period followed by test substance dosing and post-dosing periods with observations and testing. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 10-15 mice/rats per group, at least two repeated testing procedures (before, and after treatment). Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
Forced Swim Test
The forced swim test...
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- A standard design for mice includes 7 days of acclimatization period followed by test substance dosing and post-dosing periods with observations, and the day of testing. The study design includes at least 3 groups of mice (tested compound, reference compound (eg Imipramine, 20 mg/kg IP – Drug 1 at Figure above), and vehicle-treated), 10-15 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM), or noninvasive (PO, IN, or with food/drink) routes of drug administration. In the standard mice design, the transparent cylindrical glass containers (H x D: 30.0 x 19.0 cm) are filled with tap water (24ºC) to a level of 18 cm from the bottom. The duration of one FST session is 6 minutes, divided by the pre-test (first 2 minutes) and test (last 4 minutes) time. Immobility time evaluation is performed by analyzing only the test time of the session (the last 4 minutes of the session) using the video recordings. For experiments in rats, the standard design comprises 7 days of acclimatization period followed by the pre-test swimming session, substance dosing according to treatment schedule with observation period, test swimming session with the video recording. The study includes 3 groups of rats (tested compound, reference compound, and vehicle-treated) with the requested amount of animals per group. For the swimming sessions, the transparent cylindrical glass containers (H x D – 50.0 x 25.0 cm) is filled with tap water (22º – 24ºC) to a level of 30 cm or more from the bottom to prevent animals touch the cylinder’s bottom by their limbs. The duration of the pre-test is 15 minutes and the test is 5 minutes. The swimming patterns are assessed by scoring each 5 seconds period of the video stream test session
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Morris/Radial Water Maze Test
The Morris Water Maze is a...
- Method Advantages: the aversive nature of the water motivates the animal to explore the territory and rapid learning easy measuring of parameters no requirement for food deprivation less risk of potential confounding scent cues both working and reference memory can be simultaneously tested in one experiment.
- Method Limitations:
- The animal’s ability to swim and physical condition or endurance can impact the results.
- The act of being immersed in water and forced to swim can induce stress that may alter the outcomes of each repeated trial.
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- A standard study design includes 7 days of acclimatization period, 2-5 days of animal training; a single or repeated dose treatment period with observations, and the days of testing. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 15-20 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
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Nociception Assays:
Hot Plate Test
The hot plate test allows you to explore the pain response caused by heat in rodents, as well as the influence of the centrally acting analgesic on this process. Peripherally acting drugs are ineffective in this test.
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The Hotplate test was carried out using IITC hot plate analgesia meter (Life Science, USA). The smooth metal surface was heated to 56±0,5 °C. The response was measured before and 15, 30, 45, and 60 min after IV injection of Vehicle (Saline) or test compound (10 mg/kg) (Mean ± SEM). Latency was measured until the first behavioral sign of nociception (limb fluttering, licking, withdrawal from the surface or animal jumping). The cut-off time for latency in the hotplate test was set at 20 s.
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- A standard study design includes 7 days of acclimatization period; single treatment and testing. The study design includes at least 3 groups of mice (tested compound, reference compound, and vehicle-treated groups), 5-10 mice per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (PO, IN, or with food/drink) routes of drug administration.
Tail Flick Test
- The tail-flick test is the most frequently used test for nociception in animals. Unlike the Hot plate test, the tail-flick test also allows local anesthetics to be tested. During the test, radiant heat applies to the rodent’s tail, and time is measured until the animal moves the tail away because of pain caused by the heat beam. The test set for the measurement of nociceptive sensitivity of animals for the analgesic properties of substance assessment. Time increase before avoidance response to the thermal stimulus reveals as analgesic action.
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Mice were treated with Vehicle (glycerol), or Benzocaine (12% benzocaine solution in glycerol) – 30 ul of solution were applied at the tail center (2 cm) for 5 min. The level of analgesia for each mouse was measured 5, 15, 30, 60, and 120 min after application of the test compound using analgesia meter (Columbus Instruments, USA) with the intensity of thermal stimulus adjusted to a baseline tail flick latency of 5.5±0.5 s. The tail exposure to infrared radiation was leveled at the site of application of test compounds (Mean ± SEM). Each measurement was performed at triplicates. The cut-off time for latency in the tail-flick test was set at 15 s.
A standard study design includes 7 days of acclimatization period; single treatment and testing. The study design includes at least 3 groups of mice (tested compound, reference compound, and vehicle-treated groups), 5-10 mice per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or noninvasive (skin application) routes of drug administration.
Physiological Tests:
Blood pressure (BP) in rodents
- Blood pressure is measured in the tail of the mouse or rat using volume pressure recording sensor technology with the CODA (Kent Scientific, USA) mouse/rat tail-cuff system.
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Rodents’ blood pressure is measured by a tail-cuff method using the Coda Non-invasive Blood-Pressure System. Rodents are placed in plastic holders setting on the warming platform for 10 min to reduce stress and achieve a tail temperature of 32-34°C before the testing. Systolic (Sys) and diastolic (Dia) blood pressure is recorded. Mean blood pressure and pulse pressure (PP, difference between systolic and diastolic BP) values were then calculated. Typically measurement includes a set of 5 acclimations and 10 regular cycles, and the average blood pressure values are calculated taking into account accepted cycles only (M ± SEM).
A standard study design includes 7 days of acclimatization period, test substance dosing and post-dosing period with observations, and the days of testing including 3-5 days of animal training. The study design includes at least 3 groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 5-15 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes), or non-invasive (PO, IN, or with food/drink) routes of drug administration.
Grip strength testing
- The grip strength test is a non-invasive method for evaluating rodent muscle force in vivo. It allows studying the neuromuscular functions by determining the maximal peak force developed by a rodent when the operator pulls it out of a specially designed grid or bar available for both fore and hind limbs. The Grip Test is included in the Functional Observational Battery (FOB) to screen for neurobehavioral toxicity.
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The grip strength test was performed in C57BL/6J male (M) and female (F) mice of different ages using a device 47200 Grip Strength Meter (Ugo Basil, Italy). The test is performed using a grating for two limbs according to the device instructions. The study of the grip strength parameter is carried out at least three times for each animal. The average value of the force obtained in gf units (gram force), which is then converted into Newton using the formula: A ∙ 0.00980665 = N, where A is the average value of the force in gf, N – average value of the force in Newton. The reproducibility of the method for mice is 16.5% (Mean ± SEM).
A standard design includes 7 days of acclimatization followed by test substance dosing and post-dosing periods with observations and testing. The study design includes at least three groups of mice/rats (tested compound, reference compound, and vehicle-treated groups), 10-15 mice/rats per group. Treatment procedures include invasive (by IV, IP, SC, and IM routes) or non-invasive (PO, IN, or with food/drink) routes of drug administration.