Tail Vein

Using hydrodynamic tail vein injections, the investigators delivered a plasmid encoding Streptococcus pyogenes Cas9 (SpCas9) and a guide RNA targeting the site of the pathogenic Fah mutation, along with a single-strand DNA oligonucleotide with the wild-type Fah sequence to serve as the HDR repair template, into the livers of HT1 mice.

From: Genome Editing , 2021

Rodents model for toxicity testing and biomarkers

Shayne C. Gad , in Biomarkers in Toxicology, 2014

Tail vein infusions

Tail vein infusions are convenient because catheter placement can be accomplished without anesthesia. A 23-gauge or smaller needle connected to an extension set is inserted into the tail. The needle and extension set is then secured to the tail with tape. The extension set is attached to a syringe that is placed on a pump and the test compound can be infused. The tail may be taped to a wooden stick or tongue depressor to further protect the needle from being dislodged. Over-the-needle catheters are also commercially available and offer the advantage that the needle is removed once the catheter is placed in the vein and may help to prevent further penetration of the vein wall and subsequent perivascular dosing ( Rhodes and Patterson, 1979). Advantages that this technique has over permanent indwelling catheters are that the catheter is removed following dosing and will not become occluded and the animal does not have to undergo anesthesia and a surgical procedure to place the catheter. Permanent catheters have a tendency over time to develop a fibrin flap or become clotted, thus losing patency. A major disadvantage is that the animals have to be restrained during the infusion, which may cause stress and alter the results of the study. When using this technique, the duration of the infusion should be limited so that the length of time the animal is restrained is limited.

An alternative technique using the lateral tail vein involves placing a catheter in the vein and wrapping the tail in a similar manner as previously described, and then a lightweight protective cover attached to a tether system is placed around the tail to hold the catheter or needle in place.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780124046306000026

Collection of Body Fluids

Katsuhiro Fukuta , in The Laboratory Mouse (Second Edition), 2012

Bleeding from the tail vein

Tail veins are frequently used for collection of venous blood and intravenous injections. In the tail there are three veins and one artery: paired lateral caudal veins, an unpaired dorsal caudal vein and the ventral caudal artery ( Figure 5.3.8). As the tail veins are thicker than the dorsal metatarsal and medial saphenous veins, a larger quantity of venous blood can be obtained. The lateral caudal veins are preferentially used for both blood collection and intravenous injection. However, the tail should also be warmed up to dilate the blood vessel before bleeding. The mouse is warmed up as before or its tail is soaked directly in warm water at 40   °C. The mouse need not be anaesthetized, if the assistant holds the mouse, or a restraining device is used. The device in Figure 5.3.6 is used for both hindlimb and caudal veins. For the tail vein only, another restraining device is preferably used without anaesthesia (Figure 5.3.9a). The investigator should grip near the tip of the tail of the restrained mouse with the left hand, and locate the veins running down both sides of the tail. Clean from one third to one half of the tail from its end with alcohol-soaked cotton, then wipe dry with sterile gauze. The vessel is then pierced with a needle (18–21G) or incised with a razor blade. A blood droplet will accumulate on the tail which can then be collected using a glass capillary tube as before. Two or three capillary tubes are filled, giving a total of 200–300   μL of venous blood. After bleeding the incision site should be pressed with sterile gauze until the bleeding stops.

Figure 5.3.8. Schematic drawing of a transverse section of mouse tail showing the location of the dorsal caudal and lateral caudal veins and ventral caudal artery.

Paired lateral caudal veins are used for venous blood collection and intravenous injections.

Figure 5.3.9. Another restraining device composed of transparent acrylic resin.

(a) Only the tail is free for operation. (b) A winged needle is inserted into the lateral caudal vein to collect blood.

Larger volumes can be obtained in this procedure, i.e. 0.5–1.0   mL [2] or 1.0–1.5   mL [10]. However, removal of this volume of blood will cause severe hypovolaemic shock or death of the animals.

A winged intravenous drip needle is used to collect blood in laboratory animals [11]. In mice a 25G winged needle is inserted into the lateral caudal vein. A disposable 1   mL plastic syringe is connected to the vinyl tube of the winged needle and the plunger is withdrawn, so that blood enters the vinyl tube (Figure 5.3.9b). Then the blood within the tube is transferred into a microhaematocrit capillary tube by depressing the plunger. This procedure collects an adequate volume of blood (50–100   μL) and can be repeated with little damage to the blood vessel.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780123820082000313

Experimental Modeling and Research Methodology

Michael A. Koch , in The Laboratory Rat (Second Edition), 2006

b. INCISION OF LATERAL TAIL VEIN.

The tail veins of the rat can be incised to collect blood. In one report, the vein is incised longitudinally after the area is coated with petroleum jelly (Nerenberg and Zedler, 1975). Blood is collected after the tail is placed in a vacuum device fashioned from a test tube and other common laboratory materials. More recently, Fluttert et al. (2000) describes an alternative technique where rats are loosely restrained within a towel and a 2-mm incision is made in a tail vein. A drop of blood forms at the site and is collected in capillary tubes. The tail is gently stroked to facilitate blood flow. The authors consider the technique to be easy to learn and "animal friendly." However, serum enzymes such as creatine kinase were not evaluated to determine the extent of tissue trauma from this procedure. Either of these techniques can be used for serial collections since the scab that forms at the site can be easily removed.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780120749034500212

Functional Glycomics

Shingo Hatakeyama , ... Chikara Ohyama , in Methods in Enzymology, 2010

6 IV Injection into the Tail Vein

For tail vein injection, mice should be older than 6 weeks of age because at younger ages the vessel is not thick enough for injection. The injection site should be cleaned and disinfected before the operator attempts to enter the vessel.

The most important part of the procedure is the method of holding the mouse because injection needs accurate manipulation of the needle. Several restraint devices are available and these are useful for holding mice for longer periods of time (Fig. 23.1B).

Three vessels are visible on the back of the mouse's tail: a central artery with a vein on each side. You can easily distinguish an artery from a vein by the branch vessels. Branch vessels extend from the artery to the veins (Fig. 23.2A, B). Avascularization using a soft tube (Fig. 23.2C) or soaking the tail in warm water will raise the vein and make injection easier.

Figure 23.2. Tail vessels of the mouse. (A) Scheme of tail vasculature. Branch vessels extend from the artery to the veins. Red arrows, artery; blue arrows, vein; green arrows, branch vessels. (B) Careful injection is necessary in BALB/c nude (nu/nu) mice because their vessels are thin and leaky. (C) Avascularization (black arrow) or soaking the tail in warm water will make injection easier. Red, blue, and green arrows are arteries, veins, and branch vessels, respectively.

Once the mouse is safely held in the restrainer, the tail is pulled to straighten it. The mouse is monitored during the procedure by observing its respiratory rate and checking whether restraint is causing the animal any distress. The best syringe for tail vein injection is the one for insulin injection with a 28-gauge needle. The needle is bent at an angle of 30–50°. The total volume recommended for an IV injection is 100   μl. The needle is placed on the surface almost parallel to the vein and inserted carefully (Fig. 23.3A–C ). A common reason for misinjection is penetration caused by excessively deep insertion, because the vessel wall is located just beneath the skin surface. Once the needle tip is under the skin, it is very important to pull back the syringe slightly during insertion to confirm the blood will flow back (Fig. 23.3D), and then start the injection without moving the needle tip. The procedure for IV injection into the tail vein requires careful handling of the mouse and needle. Repeated practice is essential for success with this technique.

Figure 23.3. Procedure for tail vein injection. (A) Proper handling of the syringe is essential for successful injection. The outer tube is grasped by the first and second fingers. The third finger is placed under the inner cylinder. Place the needle on the surface of the tail in parallel (B) and insert it carefully (C). Once the needle tip is under the skin, pull back the syringe slightly during insertion to confirm that blood will flow back (D, arrow).

In the lung colonization assay, tumor cells (1   ×   105 to 5   ×   106 cells) in 100   μl serum-free medium are injected through the tail vein. The number of cells injected will depend on the malignant potential of the cell line. Numbers of cells commonly used for IV injections are summarized in Table 23.2.

Table 23.2. Standard numbers of cells for injection or inoculation in mice

Cell line Methods Cells/μl Periods
B16F1 IV injection 1   ×   105/100   μl 2–3 weeks
MeWo IV injection 5   ×   106/100   μl 2–3 weeks
JKT-1 IV injection 2   ×   106/100   μl 4 weeks
JKT-1 Testicular inoculation 2   ×   106/50–100   μl 3–4 weeks
PC3 Prostate inoculation 2   ×   106/20   μl 4–7 weeks
LNCaP Prostate inoculation 2   ×   106/20   μl 4 weeks
MBT-2 SC inoculation 2   ×   105/100 μl 10 days
LNCaP SC inoculation 5   ×   106/100   μl (with matrigel) 12 weeks
MDA-MB-231 MFP inoculation 1   ×   106/100   μl 30–35 days
B16F10 FP inoculation 2   ×   106/20   μl 10 days
B16F1 FP inoculation 4   ×   105/20   μl 18–21 days

Ohyama et al. (1999) injected mouse melanoma B16F1 cells stably transfected with α1,3-fucosyltransferase III (FTIII) to express sialyl Lewis X structures into the tail vein and evaluated lung tumor nodules 2–3 weeks later (Fig. 23.4). When injected to C57BL/6 mice, cells expressing moderate amounts of sialyl Lewis X (B16-FTIII-M) produced a significantly greater number of lung tumor foci than sialyl Lewis X-negative B16 cells (B16-FTIII-N). In contrast, cells expressing large amounts of sialyl Lewis X (B16-FTIII-H) produced few lung tumor foci. These results may seem to be paradoxical, because it has been postulated based on the in vitro experiments that sialyl Lewis X expression correlates with metastatic potential due to its high affinity to E-selectin.

Figure 23.4. Tumor formation in the lung. Mouse melanoma B16F1 cells were stably transfected with α1,3-fucosyltransferase III (FTIII) to express sialyl Lewis X structures. Transfected B16F1 cells (B16-FTIII cells) were separated by cell sorting into three groups based on the expression level of sialyl Lewis X (sLeX negative, moderately positive, and highly positive). When transfected cells (1   ×   105/100   μl) were injected to C57BL/6 mice, cells expressing moderate amounts of sialyl Lewis X (B16-FTIII-M) produced a significantly greater number of lung tumor foci than sialyl Lewis X-negative B16 cells (B16-FTIII-N). In contrast, cells expressing large amounts of sialyl Lewis X (B16-FTIII-H) produced few lung tumor foci. When injected to C57BL/6 mice that had been depleted of NK cells using anti-asialo-GM1 antibody, B16-FTIII-H cells that were highly positive for sialyl Lewis X produced large numbers of lung tumor nodules.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/S0076687910790236

Gene Therapy in Oncology

James E. Talmadge , Kenneth H. Cowan , in Abeloff's Clinical Oncology (Sixth Edition), 2020

Direct DNA Injection/Transduction

One form of nonviral gene delivery is the use of purified DNA plasmids. The transgene expression is low after intramuscular or intratumoral injection; however, high levels are observed if hydrodynamic injection is used. 67,68 Naked DNA injection is typically performed as an intramuscular or intratumoral injection. Despite the simplicity of this approach, transfection efficiency is low and results in limited expression. Various formulations, including lipid or pluronic formulations, and incorporation into nanoparticles or liposomes, have been used to improve transduction efficacy and gene expression. 69–71

Nonviral liposomal delivery systems can be intravenously injected with limited vector-associated toxicity, but with transgene expression, especially in the lungs. 72 Tumor targeting using tumor specific promoters, ligandation of receptors to the liposome surface, and pegylation of liposomes have all been studied. 73–79 Although some degree of tumor targeting has been observed with these delivery systems, the level of transgene expression is generally low. Studies have revealed that liposome/DNA complexes can also elicit an inflammatory response when injected systemically, resulting in suppression of transgene expression. 80–82 Furthermore, failure to achieve increased or sustained gene expression after repeated injections has been a major obstacle in the development of liposomes. 81,83 It has been shown that cationic liposome (DOTAP:cholesterol or DOTAP:Chol)/DNA complexes can achieve effective levels of transgene expression in tumor-bearing lungs and, when injected intravenously, can achieve levels sufficient to cure immunocompetent mice with disseminated experimental metastases. 84 Furthermore, repeated daily injections can result in a dose-dependent increase in transgene expression in tumor-bearing lungs. 85

Hydrodynamic Gene Delivery

Hydrodynamic tail vein plasmid delivery results in high levels of transgene expression in the livers of rodents. 86 Lower levels of transgene expression (100- to 1000-fold) are found in the spleen, heart, kidneys, and lungs. This simple nonviral gene transfer procedure entails the rapid delivery of naked plasmid DNA in a relatively large volume of physiologic saline. 67 In a typical mouse, weighing 20 g, the plasmid is delivered in a total volume of 2.0 mL over a period of 5 to 7 seconds.

Liposomes and Virosomes

In their most basic form, liposomes consist of two lipid species: a cationic amphiphile and a neutral phospholipid. 85,87 Liposomes spontaneously bind to and condense DNA to form complexes that have a high affinity for the plasma membranes of cells, resulting in the uptake of liposomes to the cytoplasm by endocytosis. Many variations of this approach are used, resulting in varying levels of gene expression. Unfortunately, liposome-facilitated gene delivery is relatively ineffectual in vivo. Some of the advantages of viral delivery vectors have been combined with the safety and "simplicity" of the liposome to produce fusogenic virosomes. 87 Virosomes are engineered by complexing the membrane fusion proteins with liposomes that have already encapsulated plasmid DNA. The inherent ability of the viral proteins in virosomes to fuse with cell membranes results in the efficient introduction of DNA to the target cell, providing improved gene expression. Viral vectors have limitations based on the size of transgene that can be incorporated; in contrast, no such limit exists for virosome or liposome technology (at least in theory).

Ballistic Delivery (Gene Gun)

Ballistic delivery, a physical method of gene delivery, uses microcarriers (usually gold particles) coated with DNA and "fired" with an explosive or gas-powered ballistic device called a gene gun. 88–90 Once the particles are inside the target cell, the DNA is slowly released from the microcarriers, transcribed, and translated. This application has been used extensively in animal models, but its clinical use is restricted to exposable surfaces or ex vivo transduction because the particles do not penetrate tissues deeply. 91

Nanoparticles

Novel polymeric delivery systems (e.g., nanospheres) are potentially useful because the smaller the size of the condensed DNA particles, the better the in vivo diffusion to target T cells and trafficking within cells. 92–94 Individual plasmid molecules can be condensed into a nanoparticles by using detergents. Nanoparticle-based gene delivery targeting the neovasculature by means of an integrin-targeting ligand has been shown to result in tumor regression. 95 Nonetheless, the size of the transgene that can be delivered is limited, and the primary focus has been on small interfering RNA (siRNA) delivery, which is discussed in a following section.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780323476744000293

Infusion Toxicology and Techniques

J.C. Resendez , D. Rehagen , in A Comprehensive Guide to Toxicology in Nonclinical Drug Development (Second Edition), 2017

Methods of Restraint and Acclimation

The lateral tail veins provide options for continuous intravenous dose administration, allowing short-term access to peripheral vessels through the skin. Because the peripheral catheter is positioned on the animal's tail, the animal must be restrained during the dose administration to protect the catheter. Typically, rats are mechanically restrained for no more than 1–2  h per dose. Mechanical restraint devices result in minimal stress to the animal while allowing easy and unhindered access to the tail. Short intermittent, peripheral infusions will require appropriate acclimation to the restraint device. Typically, the animals are acclimated to the restraint process for incremental periods up to 1   h in duration. For example, a group of rats scheduled for a 1   h infusion may be acclimated on three separate occasions for incremental durations of 15   min, 30   min, and 1   h during the pretreatment period.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780128036204000219

Common Technical Procedures in Rodents

K.L. Stewart , in Principles of Animal Research for Graduate and Undergraduate Students, 2017

8.3.1.3 Tail Vein

The distal tail vein procedure can be used to collect blood from mice and rats. The distal end of the tail is cleaned with warm water. Alcohol should not be used as it will cause vasoconstriction. Warming the tail before the procedure with a warm wash cloth or a heating pad will increase blood flow. The distal tip of the tail is amputated with a sterile surgical blade or a sharp pair of sterile surgical scissors. However, no more than 1  mm of tail tissue should be removed from a mouse or 2   mm from a rat (NIH.gov, 2010). The blood collection tube is positioned under the clipped part of the tail. Stroking the tail or squeezing the tail from the base to the distal end can increase blood flow but can also increase the contamination of the sample with other cells and other tissue products. Once sufficient blood has been collected, direct pressure is applied to the tip of the tail using a cotton ball for 20–30   s until hemostasis is confirmed. Repeated samples can be taken by the gentle removal of the clot at the end of the tail.

The lateral tail vein procedure is also used to collect blood from mice and rats. The animal is prepared as described for the distal vein collection. The tail is held between the forefinger and thumb to stabilize it. Gentle pressure is placed on the tail. A needle is used to pierce the skin, entering at a very shallow angle, almost parallel to the tail. The needle is advanced until a "flash" of blood is seen in the hub of the needle. The blood must be aspirated slowly to avoid collapsing the vessel. Once the sample is collected, the needle is removed and direct pressure is applied over the venipuncture site for 20–30   s until hemostasis has been established.

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780128021514000086

Clinical Biochemistry and Hematology

Ida M. Washington , Gerald Van Hoosier , in The Laboratory Rabbit, Guinea Pig, Hamster, and Other Rodents, 2012

Blood

Cotton rats have tail veins that are not accessible for blood sampling, although blood samples can be obtained from the cut tip of the tail without anesthesia ( Dolyak and Leone, 1953). A blood sample of up to 500   µl (per 100-g animal) may be collected from the retro-orbital venous sinus of an anesthetized cotton rat (Prince, 1994; Webb et al., 2003). Blood flow may be increased by applying pressure to the ipsilateral jugular vein (Prince, 1994). Blood sampling directly from the heart of anesthetized cotton rats has also been reported (Dolyak and Leone, 1953).

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780123809209000031

Hydrogen Peroxide and Cell Signaling, Part C

Juliano L. Sartoretto , ... Thomas Michel , in Methods in Enzymology, 2013

6 Imaging Intracellular H2O2 in Cardiac Myocytes and Endothelial Cells Expressing HyPer2

Lentiviral infection by mouse tail vein injection of the HyPer2 lentivirus leads to a heterogeneous level of HyPer2 expression in cardiac myocytes. In a typical cardiac myocyte preparation following HyPer2 tail vein injection, only approximately 5% of the myocytes express detectable HyPer2. This low proportion of HyPer2-positive cells is further confounded by the high background fluorescence present in cardiac myocytes. The intensity level of this background fluorescence has to be determined first; only cardiac myocytes showing at least twice this fluorescence level are selected for imaging. Following initiation of the live cell imaging protocol, it is important to wait until the baseline signal stabilizes: the first ~  30   s of acquired data usually cannot be analyzed because of an optical artifact that arises from the initial photoconversion of YFP (seen as a rapid drop in fluorescence), as has been previously described (Aoki & Matsuda, 2009). The individual imaging conditions (e.g., exposure time and choice of neutral density filters) depend on the specific microscope setup being used. The paper by Pase et al. (2012) contains an extensive description of these considerations. Figure 4.4 shows imaging results obtained in cardiac myocytes isolated from mice after tail vein injection with the HyPer2 lentivirus (Sartoretto et al., 2011). Changes in cell-derived fluorescence were analyzed after treating the cells with hydrogen peroxide (H2O2, 10   μM), angiotensin-II (ANG-II, 500   nM), or isoproterenol (ISO, 100   nM). HyPer2 fluorescence increases after the addition of H2O2, serving as a key positive control. ANG-II also promotes a significant increase in HyPer2 fluorescence, but there was no significant increase in HyPer2 fluorescence following the addition of ISO (Fig. 4.4). Both of these agonists promote NOS activation (Sartoretto et al., 2009, 2011), but H2O2 plays a differential role in receptor signaling: ANG-II-promoted eNOS activation is dependent on H2O2, whereas beta adrenergic receptor activation of nNOS is independent of changes in intracellular H2O2. These and similar experimental approaches can be used to probe H2O2-dependent signaling pathways in cardiac myocytes and other terminally differentiated cells.

Figure 4.4. Detection of H2O2 in cardiac myocytes isolated from mice infected with lentivirus expressing the HyPer2 biosensor. Adult mice were injected via tail vein with lentivirus expressing the HyPer2 H2O2 biosensor (~   109  pfu); 2 weeks later the mice were euthanized, and cardiac myocytes were isolated and analyzed. The bar graph shows pooled data from three independent experiments, in which the H2O2 response is quantitated as the slope of the fluorescence signal in arbitrary units (AU) measured between t  =   0 and t  =   5   min after the addition of 10   μM of H2O2, 500   nM of ANG-II (angiotensin-II), or 100   nM of ISO (isoproterenol). *p  <   0.05 compared to PBS-treated cells. Also presented are representative HyPer2 images shown in isolated cardiac myocytes treated as displayed. The HyPer2 H2O2 image is determined as the YFP500/YFP420 excitation ratio; the gray scale is adjusted to improve contrast.

 Adapted with modifications from Sartoretto et al. (2011).

Read full chapter

URL:

https://www.sciencedirect.com/science/article/pii/B9780124058811000045