IMPC Pipeline Schedule

Unrestricted

Welfare Observations IMPC_WEL_001 (v.1.0)

The Welfare Observations procedure is used for the recording of welfare issues as and when they occur.

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Body Weight IMPC_BWT_001 (v.1.3)

The body weight test measures the weight of the mouse in a time series, allowing monitoring of its evolution; also, it is required in many other procedures.

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Experimental design IMPC_EXD_002 (v.2.0)

experimental_design

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Previous Versions:(v.1.0)

Adult LacZ IMPC_ALZ_001 (v.1.9)

To stain adult mutant and control tissues for bacterial beta-galactosidase (LacZ) activity in order to assess the adult organ, tissue, substructure and cell type of gene expression of IKMC alleles utilizing a LacZ reporter. Description: Adult mouse tissues are scored for presence of LacZ staining which is distinct from either nonspecific staining observed in wildtype control mice, or is too faint to score as present. Intensity of staining is not required but can be reported. Anatomical terms to localize staining to the organ, structure, substructure, or cell type will use standard mouse anatomy ontology terms.

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Fertility of Homozygous Knock-out Mice IMPC_FER_001 (v.1.1)

To assess the fertility of homozygous knockout mice.

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Viability Primary Screen IMPC_VIA_002 (v.2.0)

Assess the viability of mutant mice of each sex and zygosity.

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Previous Versions:(v.1.0)

Housing and Husbandry IMPC_HOU_002 (v.2.0)

Housing_and_husbandry

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Previous Versions:(v.1.0)

E9.5 and Younger

Gross Morphology Embryo E9.5 IMPC_GEL_003 (v.3.0)

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Previous Versions:(v.2.0)(v.1.0)

Viability E9.5 Secondary Screen IMPC_EVL_001 (v.1.0)

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OPT E9.5 IMPC_EOL_002 (v.2.0)

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Previous Versions:(v.1.0)

MicroCT E9.5 IMPC_EML_002 (v.2.0)

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Previous Versions:(v.1.0)

Gross Morphology Placenta E9.5 IMPC_GPL_001 (v.1.0)

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E12.5

Gross Morphology Embryo E12.5 IMPC_GEM_003 (v.3.0)

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Previous Versions:(v.2.0)(v.1.0)

E15.5

Viability E14.5-E15.5 Secondary Screen IMPC_EVO_001 (v.1.0)

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Gross Morphology Placenta E14.5-E15.5 IMPC_GPO_001 (v.1.0)

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MicroCT E14.5-E15.5 Analysis IMPC_MAA_001 (v.1.0)

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MicroCT E14.5-E15.5 IMPC_EMO_002 (v.2.0)

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Previous Versions:(v.1.0)

Gross Morphology Embryo E14.5-E15.5 IMPC_GEO_003 (v.3.0)

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Previous Versions:(v.2.0)(v.1.0)

E18.5 and Older

Viability E18.5 Secondary Screen IMPC_EVP_001 (v.1.0)

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MicroCT E18.5 IMPC_EMA_002 (v.2.0)

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Previous Versions:(v.1.0)

Gross Morphology Embryo E18.5 IMPC_GEP_003 (v.3.0)

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Previous Versions:(v.2.0)(v.1.0)

Gross Morphology Placenta E18.5 IMPC_GPP_001 (v.1.0)

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Week 9

Open Field IMPC_OFD_001 (v.1.1)

The Open Field test is used to assess anxiety and exploratory behaviors. It is based on the natural tendency of an animal to explore and to protect itself using avoidance which translates to a normal animal spending more time in the periphery of the Open Field arena than in the center (the most anxiogenic area).

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Grip Strength IMPC_GRS_001 (v.1.1)

The grip strength test is used to measure the neuromuscular function as maximal muscle strength of forelimbs and combined forelimbs and hind limbs. These are assessed by the grasping applied by the mouse on a grid that is connected to a sensor. Three trials are carried out in succession measuring forelimb-strength only, followed by three successive trials measuring the combined forelimb/hindlimb grip strength. Ontological description: MP:0001515 - abnormal grip strength.

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Combined SHIRPA and Dysmorphology IMPC_CSD_003 (v.3.9)

SHIRPA and dysmorphology were originally always separate assessments. However they have recently been combined as assessments, so that they take place at the same time.The purpose of the assessments is to examine mice for obvious physical characteristics, behaviors and morphological abnormalities.Descriptions include abnormal locomotion/appearance/behavior/reflex reactions.

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Previous Versions:(v.2.7)(v.1.11)

Week 10

Acoustic Startle and Pre-pulse Inhibition (PPI) IMPC_ACS_003 (v.3.4)

The acoustic startle response is characterized by an exaggerated flinching response to an unexpected strong auditory stimulus (pre-pulse). This response can be attenuated when it is preceded by a weaker stimulus (pre-pulse) and is the principle underlying pre-pulse inhibition (PPI). PPI has been described in numerous species, including mice and humans and provides an operational measure of sensorimotor gating reflecting the ability of an animal to successfully integrate and inhibit sensory information. Several clinical studies have shown that a number of human disorders have impaired PPI including: schizophrenia, Huntington’s disease, fragile X syndrome, and autism. The acoustic startle and PPI paradigm is therefore largely used to assess sensorimotor gating and the effects of a number of treatment modalities such as putative anti-psychotics, and to explore genetic and neurobiological mechanisms underlying behaviors of relevance to psychosis (Geyer, 1999; Ouagazzal et al., 2001).Ontological description: MP:0002067 - abnormal sensory capabilities/reflexes/nociception.

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Previous Versions:(v.2.4)(v.1.5)

Light-Dark Test IMPC_LDT_001 (v.1.1)

Light-Dark test monitors the anxiety-related behaviour of mice

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Week 11

Fear Conditioning IMPC_FEA_001 (v.1.2)

Fear Conditioning test assesses aversive learning and memory

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Indirect Calorimetry IMPC_CAL_003 (v.3.5)

Indirect calorimetry provides detailed information on the energy metabolism of mutant mice. Energy expenditure is evaluated through indirect calorimetry by measuring oxygen consumption with an open flow respirometric system. CO2 and O2 sensors measure the difference in CO2 and O2 concentrations in air volumes flowing through control or animal cages. The amount of oxygen consumed over a given period of time can thus be calculated, as far as the air flow through the cage is known. Data are expressed as ml O2 h-1animal-1. The system also monitors CO2 production, therefore, the respiratory exchange ratio (RER) and heat production can be calculated. An activity and food and water intake monitoring system can also be integrated into the set up in order to investigate circadian pattern and behaviour.Ontological description: MP:0005266 - abnormal metabolism.

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Previous Versions:(v.2.7)(v.1.8)

Y-maze IMPC_YMZ_001 (v.1.1)

Y-maze assesses learning and memory; it can be used instead/in parallel of Fear Conditioning

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Week 12

Echo IMPC_ECH_001 (v.1.4)

To assess the functionality of the heart in order to determine the presence of a mutant phenotype.

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Electrocardiogram (ECG) IMPC_ECG_003 (v.3.0)

To provide a high throughput method to obtain Electrocardiograms in a conscious mouse or an anesthetized mouse.

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Previous Versions:(v.2.6)(v.1.9)

Week 13

Intraperitoneal glucose tolerance test (IPGTT) IMPC_IPG_001 (v.1.1)

The glucose tolerance test measures the clearance of an intraperitoneally injected glucose load from the body. It is used to detect disturbances in glucose metabolism that can be linked to human conditions such as diabetes or metabolic syndrome. Animals are fasted for approximately 16 hours (unless specified otherwise), fasted blood glucose levels are determined before a solution of glucose is administered by intra-peritoneal (IP) injection. Subsequently, the blood glucose level is measured at different time points during the following 2 hours.Ontological description: MP:0005559 - increased circulating glucose level, MP:0005560 - decreased circulating glucose level, MP:0005293 - impaired glucose tolerance, MP:0005292 - improved glucose tolerance, MP:0005291 abnormal glucose tolerance, MP:0000188 - abnormal circulating glucose level.

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Challenge Whole Body Plethysmography IMPC_CHL_001 (v.1.1)

The purpose of this procedure is to record the respiratory function of mice, when submitted to methacholine or hypoxia challenge. Other similar protocols, for allergen sensitization and for LPS challenges, will also be available. Ontological description: MP:0002327 - abnormal respiratory function

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Week 14

Auditory Brain Stem Response IMPC_ABR_002 (v.2.5)

Auditory brainstem response test determines hearing sensitivity and other physiological parameters using evoked potential recordings in anesthetized mice.Ontological description: MP:0004738 - abnormal brainstem auditory evoked potential.

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Previous Versions:(v.1.7)

X-ray IMPC_XRY_001 (v.1.9)

Construct and analyse digital X-ray images in immobilised mice using a Faxitron X-Ray system or NTB digital X-ray scanner.

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Body Composition (DEXA lean/fat) IMPC_DXA_001 (v.1.8)

Measure bone mineral content and density as well as body composition in mice using the DEXA (Dual Energy X-ray Absorptiometry) analyser.

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Week 15

Eye Morphology IMPC_EYE_003 (v.3.0)

To detect abnormalities in eye morphology.

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Previous Versions:(v.2.8)(v.1.9)

Week 16

Tissue Embedding and Block Banking IMPC_BLK_001 (v.1.9)

Collect and fix a standard list of tissues from the complete necropsy (see IMPC Gross Pathology & Tissue Collection SOP)Trim the fixed tissues into cassettes for processingEmbed the tissues in paraffin in a standard orientation

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Clinical Chemistry IMPC_CBC_003 (v.3.9)

Clinical chemistry determines biochemical parameters in plasma including enzymatic activity, specific substrates and electrolytes. Ontological description: MP:0001545 – blood physiology abnormalities.

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Previous Versions:(v.2.1)(v.1.12)

Gross Pathology and Tissue Collection IMPC_PAT_002 (v.2.9)

To perform a complete necropsy to detect and record abnormal external findings and macroscopic alterations in internal and external organs, record body and heart weights (see IMPC Heart Weight SOP), and collect a standardized list of tissues for fixation with or without further processing (see non-mandatory IMPC Tissue Embedding & Block Banking SOP).

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Previous Versions:(v.1.9)

Histopathology IMPC_HIS_003 (v.3.0)

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Previous Versions:(v.2.6)(v.1.9)

Hematology IMPC_HEM_002 (v.2.9)

Hematological assessment of blood determines blood cell counts (white blood cells, red blood cells, hemoglobin, and platelets) and additional hematological parameters (hematocrit, mean cell volume, mean corpuscular hemoglobin, mean cell hemoglobin concentration) can be derived using these indices. These tests will indicate abnormalities in the production of blood and its components (blood cells and hemoglobin) as well as in the associated blood-forming organs. Ontological description: MP:0002429 - abnormal blood cell morphology/development.

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Previous Versions:(v.1.11)

Organ Weight IMPC_OWT_001 (v.1.4)

Organ weights from different tissues collected for IMPC

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Heart Weight IMPC_HWT_001 (v.1.9)

To evaluate cardiac size using heart weight and body weight.

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Insulin Blood Level IMPC_INS_003 (v.3.2)

The insulin concentration in the blood is an important indicator of diabetes.Ontological description: abnormal circulating insulin level [MP:0001560]; increased circulating insulin level [MP:0002079]; decreased circulating insulin level [MP:0002727].

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Previous Versions:(v.2.3)(v.1.3)

Immunophenotyping IMPC_IMM_002 (v.2.0)

Immunophenotyping of immune system cells

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Previous Versions:(v.1.8)

Formalin Test IMPC_FOR_001 (v.1.0)

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Phenotyping Protocols for Pipeline: IMPC Pipeline IMPC_001

Unrestricted

Welfare Observations IMPC_WEL_001 (v.1.0)

Body Weight IMPC_BWT_001 (v.1.3)

Experimental design IMPC_EXD_002 (v.2.0)

Adult LacZ IMPC_ALZ_001 (v.1.9)

Fertility of Homozygous Knock-out Mice IMPC_FER_001 (v.1.1)

Viability Primary Screen IMPC_VIA_002 (v.2.0)

Housing and Husbandry IMPC_HOU_002 (v.2.0)

E9.5 and Younger

Gross Morphology Embryo E9.5 IMPC_GEL_003 (v.3.0)

Viability E9.5 Secondary Screen IMPC_EVL_001 (v.1.0)

OPT E9.5 IMPC_EOL_002 (v.2.0)

MicroCT E9.5 IMPC_EML_002 (v.2.0)

Gross Morphology Placenta E9.5 IMPC_GPL_001 (v.1.0)

E12.5

Gross Morphology Embryo E12.5 IMPC_GEM_003 (v.3.0)

Embryo LacZ IMPC_ELZ_001 (v.1.1)

Gross Morphology Placenta E12.5 IMPC_GPM_001 (v.1.0)

Viability E12.5 Secondary Screen IMPC_EVM_001 (v.1.0)

E15.5

Viability E14.5-E15.5 Secondary Screen IMPC_EVO_001 (v.1.0)

Gross Morphology Placenta E14.5-E15.5 IMPC_GPO_001 (v.1.0)

MicroCT E14.5-E15.5 Analysis IMPC_MAA_001 (v.1.0)

MicroCT E14.5-E15.5 IMPC_EMO_002 (v.2.0)

Gross Morphology Embryo E14.5-E15.5 IMPC_GEO_003 (v.3.0)

E18.5 and Older

Viability E18.5 Secondary Screen IMPC_EVP_001 (v.1.0)

MicroCT E18.5 IMPC_EMA_002 (v.2.0)

Gross Morphology Embryo E18.5 IMPC_GEP_003 (v.3.0)

Gross Morphology Placenta E18.5 IMPC_GPP_001 (v.1.0)

Week 9

Open Field IMPC_OFD_001 (v.1.1)

Grip Strength IMPC_GRS_001 (v.1.1)

Combined SHIRPA and Dysmorphology IMPC_CSD_003 (v.3.9)

Week 10

Acoustic Startle and Pre-pulse Inhibition (PPI) IMPC_ACS_003 (v.3.4)

Light-Dark Test IMPC_LDT_001 (v.1.1)

Week 11

Fear Conditioning IMPC_FEA_001 (v.1.2)

Indirect Calorimetry IMPC_CAL_003 (v.3.5)

Y-maze IMPC_YMZ_001 (v.1.1)

Week 12

Echo IMPC_ECH_001 (v.1.4)

Electrocardiogram (ECG) IMPC_ECG_003 (v.3.0)

Week 13

Intraperitoneal glucose tolerance test (IPGTT) IMPC_IPG_001 (v.1.1)

Challenge Whole Body Plethysmography IMPC_CHL_001 (v.1.1)

Week 14

Auditory Brain Stem Response IMPC_ABR_002 (v.2.5)

X-ray IMPC_XRY_001 (v.1.9)

Body Composition (DEXA lean/fat) IMPC_DXA_001 (v.1.8)

Week 15

Eye Morphology IMPC_EYE_003 (v.3.0)

Week 16

Tissue Embedding and Block Banking IMPC_BLK_001 (v.1.9)

Clinical Chemistry IMPC_CBC_003 (v.3.9)

Gross Pathology and Tissue Collection IMPC_PAT_002 (v.2.9)

Histopathology IMPC_HIS_003 (v.3.0)

Hematology IMPC_HEM_002 (v.2.9)

Organ Weight IMPC_OWT_001 (v.1.4)

Heart Weight IMPC_HWT_001 (v.1.9)

Insulin Blood Level IMPC_INS_003 (v.3.2)

Immunophenotyping IMPC_IMM_002 (v.2.0)

Formalin Test IMPC_FOR_001 (v.1.0)