Cardiac puncture

Male CD-I mice were allowed 3 days to stabilize after arrival at the test facility. Groups of 10 mice each received 0.2 mL of the various concentrations of PGG in sterile saline by bolus intravenous (iv) injection (transthoracic cardiac puncture). A control group received 0.3 mL of sterile saline. Mice were returned to their cages, maintained on food and water ad libitum and were challenged 3-4 hours... 

Blood can be taken from fetuses at necropsy by cardiac puncture before fetal examination. It is also possible to take, from satellite litters, fetal blood or amniotic fluid during gestation (i.e., day 19 of gestation). The fetal blood is pooled by litter to obtain a sufficient plasma volume for analyzes. 

In toxicokinetic studies involving sequential animal sacrifice and tissue examination, it is critical to obtain uncontaminated organ samples. Apart from contamination by blood, suitable samples can be obtained by careful dissection and rinsing of the organs in ice-cold buffer, saline, or other appropriate solution. Blood samples themselves are obtained by cardiac puncture, and blood contamination of organ samples is minimized by careful bleeding of the animal at the time of sacrifice or, if necessary, by perfusion of the organ in question. 

By cardiac puncture, when animal survival is not required, exsan-guinations by cardiac puncture yields the maximal volume of blood ... 

Citrated plasma Kill the guinea pigs by C02-induced asphyxia. (To reduce the number of animals used, citrated plasma for future experiments may be prepared from the eosinophil donors, as in Subheading 3.1.2.) Take blood into ACD by cardiac puncture (2 mL ACD per 8 mL blood). Mix and centrifuge (2800g, 10 min, 20°C). Collect the supernatant and discard the pellet. Repeat the centrifugation and store the cell-free citrated plasma in aliquots at -20°C. 

After a predetermined time period of eosinophil accumulation, anesthetize the animals with Sagatal (60 mg/kg). Collect a blood sample (at least 3 mL) into heparin (10 IU/mL) by cardiac puncture and humanely kill the animal with an... 

Serum was obtained from blood acquired by cardiac puncture of mouse. 

Animals immunized with Freund s emulsions should be bled 7-10 days after booster injections. If the blood is taken from a vein rather than by cardiac puncture, two or three bleeds can be taken on successive days, but the animal should then be rested for 3-4 weeks before further bleeding or before boosting again if the original antiserum was not of satisfactory quality. After intravenous injection antibody levels rise and then fall more rapidly and bleeds should be collected 5-7 days after the last dose. It is often helpful to fast the animals overnight to minimize lipemia, but do not deprive them of drinking water. 

Rats dosed orally at 10 mg/kg (3 rats/ sex/timepoint) were exsanguinated by cardiac puncture while under methoxyflurane anesthesia. Blood samples were collected in heparinized syringes and kept on ice until centrifugation. Blood was spun at 2000 rpm for 10 min. Plasma was removed and stored at - 30°C until analysis. 

The method described by Hirsch et al. (H4) uses intact male rats previously maintained on a low-calcium diet for 4 days. Standard and unknown preparations are injected subcutaneously into parallel groups of test animals, and after 1 hour blood samples are collected by cardiac puncture under ether anesthesia. The plasma calcium levels are measured, and the relative potencies of the preparations are calculated by standard statistical procedures. The mean index of precision of ten consecutive assays was reported as 0.23 0.025. 

Figure 7.1 Analysis of serum IgE concentrations following exposure of mice to allergens or vehicle. Groups of mice (n=6) were exposed to 50 pi of 25 per cent TMA or 1 per cent DNCB bilaterally on the shaved flanks. Control mice received identical treatment with vehicle (AOO) alone. Seven days later, 25 pi of the same test chemical at half the application concentration used previously, or an equal volume of vehicle alone, were applied to the dorsum of both ears. Fourteen days following the initiation of exposure, mice were exsanguinated by cardiac puncture and serum prepared. Serum IgE was measured using a sandwich ELISA. Results are expressed as mean serum IgE concentration in pg. ml 1 SE (where these exceeded 0.075 pg. ml-1). A summary of six independent experiments a-f.

In conclusion, the experimental protocol recommended presently is as follows. Groups of six BALB/c strain mice are exposed topically on both shaved flanks to 50 pi of one of three concentrations of the test material or to an equal volume of vehicle alone. Additional groups of mice receive 25 per cent TMA, 1 per cent DNCB or the same volume of 4 1 acetone olive oil, the vehicle of choice for these chemicals. Seven days later mice are treated on the dorsum of both ears with 25 pi of the same chemical at half the application concentration used previously, or with the same volume of vehicle alone. Fourteen days following the initiation of exposure, mice are exsanguinated by cardiac puncture and serum prepared and stored at -20°C until analysis (Hilton et al., 1995, 1996). This protocol is illustrated diagrammatically in Figure 7.2. 

Collect as much blood as possible through cardiac puncture using a 1-in., 25-gauge needle (rrrNote 10). Transfer blood from syringe to 2mL Eppendorf tube(s) containing lmL... 

Some clotting of the blood can be prevented by coating the syringe with PBS + 2mM EDTA prior to performing the cardiac puncture. 

Cardiac puncture can be used in larger fish. Trauma in conjunction with the small size of the heart make this a less desirable alternative. The heart in most species can be approached from the caudal border of the pectoral girdle. 

Rakusan, K. and Blahitka, J., Cardiac output distribution in rats measured by injection of radioactive microspheres via cardiac puncture. Can. J. Physiol. Pharmacol, 52, 230, 1974. 

Numerous methods and data have been published for the collection of blood from rats and mice (Riley 1960 Upton and Morgan 1975 Cardy and Warner 1979 Eowler, Brown, and Elower 1980 Archer and Riley 1981 Cochetto and Bjornsson 1983 Neptun, Smith, and Irons 1985 Suber and Kodell 1985 Conybeare et al. 1988 Dameron et al. 1992 Itumi et al. 1993 Matsuzawa et al. 1994 Bernard et al. 1996 Walter 1999 Mahl et al. 2000 Nahas et al. 2000 Schnell et al. 2002). Values obtained from major blood vessels or cardiac puncture are less variable than those samples taken from the tail or retro-orbital plexus this may be in part due to contamination with tissue fluid. Potassium, total protein, and several enzymes are higher in samples collected from the tail or the retro-orbital plexus. The use of carbon dioxide and, to a lesser extent, halothane increases plasma levels of glucose, potassium, and inorganic phosphate. 

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