We would like to thank the Equine Veterinary Hospital and the Ruminants Veterinary Hospital of the College of Veterinary Medicine, University of Sao Paulo (FMVZ/USP), Sao Paulo, Brazil, FAPESP (grant 2022/11459-3, Sao Paulo Research Foundation), EMS Pharma, CNPq (grant 405254/2022-9), and &#193;gua Branca pig farming, Itu, Sao Paulo, Brazil.

This study was approved by the Ethics Committee on Animal Use in Research of the College of Veterinary Medicine and Animal Science of the University of S&#227;o Paulo, protocol number 6088030523. Nine seven-month-old gilts from the &#193;gua Branca nucleus pig farming, located in Itu city, S&#227;o Paulo state, Brazil, were used right after the second estrus detection9. The details of the reagents and the equipment used in the study are listed in the Table of Materials.
1. Animal preparation
<ol>
	<li>Fast the animals for 12 h and restrict water access for 4 h before the surgical procedure.</li>
	<li>Using a 40 mm x 12 mm needle, draw up 10 mg/kg of ketamine and 0.2 mg/kg of midazolam in the same 50 mL syringe. Attach the syringe to an IV administration set to facilitate medication administration, maintaining a safe distance from the animal and allowing it to move freely.</li>
	<li>Administer the pre-anesthetic medication via intramuscular injection into the trapezius muscle, caudal to the external ear canal.</li>
	<li>Cannulate the auricular vein with a 20 G catheter and connect it to a macrodrip administration set for intraoperative fluid therapy with 0.9% NaCl solution at a rate of 10 mL/kg/h.</li>
	<li>Induce anesthesia with 4 mg/kg of propofol intravenously (following institutionally approved protocols).</li>
	<li>Perform orotracheal intubation of the animal using a laryngoscope and a size 9 orotracheal tube with a cuff10.</li>
	<li>Maintain anesthesia with 1.5% to 2% isoflurane inhalation. Additionally, monitor anesthesia by evaluating physiological parameters such as heart rate, which should be maintained between 100 and 120 beats per minute, oxygen saturation, which should be kept between 99% and 100%, muscle relaxation, absence of movement, and reflexes evaluation. 
	NOTE: Body temperature should also be monitored and controlled, staying between 37.5-38.5 &#176;C. When necessary, in addition to isoflurane, propofol can be used to maintain control of the parameters. Fentanyl hydrochloride should be administered as an analgesic at a dose of 0.02 mg/kg every 20 min. All parameters should be recorded every 5 min.</li>
	<li>Administer perioperative analgesia with a single dose of 0.002 mg/kg of fentanyl intravenously and 4 mg/kg of tramadol hydrochloride intramuscularly into the gluteal or trapezius muscle.</li>
	<li>Place cardiac electrodes on the thoracic region for intraoperative monitoring of heart rate. Record heart rate, rectal temperature, and oxygen saturation every 5 min.</li>
</ol>
2. Surgical procedure
<ol>
	<li>Position the gilt on the surgical table in dorsal recumbency.</li>
	<li>Cover the animal&#39;s legs with procedural gloves to minimize contamination of the surgical field.</li>
	<li>Perform ventral caudal abdominal hair removal between the penultimate pair of mammary glands using a surgical clipper.</li>
	<li>Perform preoperative skin antisepsis with a chlorhexidine-impregnated antiseptic brush, making circular movements from the umbilical scar region to the last pair of mammary glands. Remove chlorhexidine with a clean, moistened compress.</li>
	<li>Ensure that the surgeon and the assistant perform surgical antisepsis and wear sterile surgical gowns and gloves for surgery. 
	NOTE: The animal should be prepared for surgery in a preparation area separate from the surgical site.</li>
	<li>Prepare the surgical table with the appropriate and previously sterilized instruments listed in the Table of Materials.</li>
	<li>Perform final surgical field antisepsis using sterile gauze and chlorhexidine antiseptic, cleaning the region in a fishbone pattern, starting at the incision site and expanding laterally.</li>
	<li>Repeat this procedure three times with chlorhexidine antiseptic, and then repeat the procedure with alcoholic chlorhexidine.</li>
	<li>Completely cover the animal with a sterile surgical drape, and using scissors, create a rectangular opening in the surgical drape, approximately 10 x 20 cm&#178;, if the drape is not fenestrated.</li>
	<li>Position the surgical drape over the penultimate pair of inguinal mammary glands. Place 4 Backhaus clamps to secure the surgical drape over the animal&#39;s skin.</li>
	<li>Incise the skin with a scalpel, creating an incision approximately 10 cm in length.</li>
	<li>Achieve hemostasis of subcutaneous vessels with hemostatic forceps and/or absorbable suture size 2.</li>
	<li>Dissect the subcutaneous tissue with fingers or with the aid of fine-pointed scissors, deepening the incision to reach the linea alba in the abdominal musculature. 
	NOTE: Palpate the midline of the abdomen to feel the linea alba as a more rigid and fibrous area; observe the separation of the rectus abdominis muscles and note the change in tissue consistency while dissecting to identify the linea alba accurately11.</li>
	<li>With Allis forceps, grasp the musculature and lift it to make a stab incision in the linea alba with a scalpel.</li>
	<li>Insert the index finger into the incision to assess the presence of adhesions. Extend the incision over the linea alba caudally and cranially with blunt/blunt or fine/blunt scissors carefully to avoid injuring other organs and underlying structures.</li>
	<li>Make an incision of sufficient length so that the surgeon&#39;s one hand can be inserted into the abdominal cavity to locate and exteriorize the uterine horns and/or ovaries. A Gosset retractor may be used to facilitate entry into the abdominal cavity and suturing of the musculature in a single layer at the end of the procedure.</li>
	<li>If one of the uterine horns is located before the ovaries, gently follow it until reaching the tip of the uterine horn and the ipsilateral ovary. During this procedure, assess the presence of fluid inside the uterus, which may compromise gestation.</li>
	<li>After locating the ovary, gently pull it and carefully expose it. Assess the presence of pre-ovulatory follicles, hemorrhagic corpora lutea, cyclic corpora lutea, and corpus albicans to evaluate if the gilt&#39;s synchronization is as expected.</li>
	<li>Gently remove the fimbriae of the uterine tube covering the ovary, evert its mucosa, and locate the ostium of the uterine tube.&#160; 
	NOTE:&#160;If the surgeon finds it difficult to identify the ostium of the fimbriae, a pocket on the ampulla of the oviduct can be done with a round-bodied suture needle, avoiding puncturing blood vessels, to insert the Tomcat catheter containing the embryos.</li>
	<li>Carefully insert a Tomcat catheter containing the embryos into the ostium of the fimbriae until reaching the ampulla region of the uterine tube.</li>
	<li>Attach a 1 mL syringe to the Tomcat catheter to push the embryo-containing fluid into the uterine tube. Cover the ovary with the fimbriae and return it to the abdominal cavity.</li>
	<li>Locate the contralateral uterine horn and ovary and repeat steps 2.17-2.21.</li>
	<li>Reposition the structures inside the abdominal cavity and add 1 L of saline solution or Ringer&#39;s lactate solution warmed (39 &#176;C) to prevent adhesions before initiating abdominorraphy.</li>
	<li>Suture the abdominal musculature with absorbable suture size 2 in &#34;X&#34; stitches (Sultan) interrupted.</li>
	<li>Approximate the subcutaneous tissue with a continuous mattress suture using absorbable suture size 2.</li>
	<li>Close the skin with size 2 nylon suture in interrupted simple stitches, surgical staples, or ethyl-cyanoacrylate adhesive. In this procedure, we opted for the use of adhesive.</li>
	<li>Clean the surgical wound with gauze soaked in hydrogen peroxide and dry with a compress to enhance the fixation of the surgical adhesive on the surgical wound. Apply rifamycin or Pearson&#39;s ointment, cover with gauze, and finally, apply surgical adhesive. 
	NOTE: The wound should remain closed with a dressing for 5 days.</li>
</ol>
3. Postoperative care
<ol>
	<li>Evaluate the animals for signs of characteristic clinical pain for at least the next 3 consecutive days, according to the scale proposed by Luna et al.12. 
	NOTE: This scale is based on different behaviors exhibited by pigs when experiencing different levels of pain, such as posture, interaction and interest in the environment, activity, appetite, and attention to the affected area. The score ranges from 0 to 3, depending on the type of behavior exhibited, for each parameter evaluated. At the end of the evaluation, the sum of all scores reflects the intensity of pain experienced by the animal, where 0 represents no pain, and 18 represents very intense pain12.</li>
	<li>Administer a single dose of 15 mg/kg of amoxicillin and 0.4 mg/kg of 2% meloxicam both intramuscularly (in the gluteal or cervical muscle) once a day for 3 days as postoperative care to prevent pain, inflammatory processes, and infection. 
	NOTE: If the pain score is equal to or greater than 6 on the pain scale used (score from 0 to 18), administer 5,000 mg/animal of sodium dipyrone intramuscularly (in the gluteal or cervical muscle) once a day in addition to the anti-inflammatory and antibiotic. Alternatively, perform analgesia as outlined in the local IACUC guidelines.</li>
	<li>On the 6th day after surgery, remove the surgical adhesive for wound cleaning with gauze soaked in 2% alcoholic chlorhexidine solution and apply topical Pearson&#39;s ointment until the 10th day, when suture removal or surgical staples removal, if used, is recommended.</li>
</ol>
4. Ultrasonographic gestational diagnosis of pregnancy
<ol>
	<li>Perform transabdominal pregnancy ultrasonographic evaluation with an ultrasound device (frequency range 4.5 to 8.5 MHz) 30 days after embryo transfer surgery.</li>
	<li>Apply ultrasound gel to the probe and position it near the inguinal abdominal region, facing medially. 
	NOTE: Visualization of rounded anechoic embryonic vesicles, due to fluid in the uterine lumen, indicates gestation. Non-visualization of intrauterine fluid, or&#160;presence of intrauterine fluid without delineation of embryonic vesicles, suggests the absence of gestation.</li>
</ol>
5. Euthanasia of the animals
NOTE: &#160;Euthanize the gilts that did not present fetuses of size and appearance corresponding to gestational age on the ultrasonographic examination.
<ol>
	<li>Using a 40 mm x 12 mm needle, draw up 15 mg/kg of ketamine and 2 mg/kg of midazolam in the same 50 mL syringe attached to an intravenous administration set, and administer intramuscularly into the trapezius muscle in the cervical region caudal to the external ear canal, as preanesthetic medication.</li>
	<li>Cannulate the auricular vein with a 20 G catheter and induce anesthesia with 4 mg/kg of propofol intravenously in a bolus until bradycardia occurs (following institutionally approved protocols).</li>
	<li>Subsequently, administer 10 to 20 mL of 19.1% potassium chloride intravenously. Confirm cardiopulmonary arrest by observing the absence of respiratory movements, pale mucous membranes, loss of corneal reflex, and absence of heartbeats by placing a stethoscope over the cardiac region.</li>
	<li>Collect any type of intrauterine content (embryonic tissue, liquid, or mucous secretions) from necropsied females13 and forward it for microbiological analysis to detect possible pathogens and also for genotypic characterization of the clones (if needed). Visually check the presence or absence of corpora lutea, ovarian cysts, abnormal secretions, and the occurrence of adhesions in the uterus, ovaries, and oviducts14.</li>
</ol>

Optimizing Oviductal Embryo Transfer in Cloned Pigs

<ol>
	<li>Lunney, J. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Importance+of+the+pig+as+a+human+biomedical+model.">Importance of the pig as a human biomedical model.</a> Sci Transl Med. 13 (621), 5758 (2021).</li><li>Xi, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetically+engineered+pigs+for+xenotransplantation:+Hopes+and+challenges.">Genetically engineered pigs for xenotransplantation: Hopes and challenges.</a> Front Cell Dev Biol. 10, 1093534 (2023).</li><li>Chen, P. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Production+of+pigs+from+porcine+embryos+generated+in+vitro.">Production of pigs from porcine embryos generated in vitro.</a> Front Anim Sci. 3, 826324 (2022).</li><li>Wieczorek, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Laparoscopic+embryo+transfer+in+pigs+-+comparison+of+different+variants+and+efficiencies+of+the+method.">Laparoscopic embryo transfer in pigs - comparison of different variants and efficiencies of the method.</a> Pol J Vet Sci. 26 (2), 295-306 (2023).</li><li>Martinez, E. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Design,+development,+and+application+of+a+non-surgical+deep+uterine+embryo+transfer+technique+in+pigs.">Design, development, and application of a non-surgical deep uterine embryo transfer technique in pigs.</a> Anim Front. 3 (4), 40-47 (2013).</li><li>Wieczorek, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+new+concept+in+minimally+invasive+embryo+transfer.">A new concept in minimally invasive embryo transfer.</a> Ann Anim Sci. 20 (4), 1289-1308 (2020).</li><li>Brussow, K. P., Torner, H., Kanitz, W., Ratky, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vitro+technologies+related+to+pig+embryo+transfer.">In vitro technologies related to pig embryo transfer.</a> Reprod Nutr Dev. 40 (5), 469-480 (2000).</li><li>Chen, P. R., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Production+of+pigs+from+porcine+embryos+generated+in+vitro.">Production of pigs from porcine embryos generated in vitro.</a> Front Anim. Sci. 3, 826324 (2022).</li><li>Langendijk, P., vanden Brand, H., Soede, N. M., Kemp, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+boar+contact+on+follicular+development+and+on+estrus+expression+after+weaning+in+primiparous+sows.">Effect of boar contact on follicular development and on estrus expression after weaning in primiparous sows.</a> Theriogenology. 54 (8), 1295-1303 (2000).</li><li>Costea, R., Ene, I., Pavel, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pig+sedation+and+anesthesia+for+medical+research.">Pig sedation and anesthesia for medical research.</a> Animals. 13, 3807 (2023).</li><li>Vierstraete, M., Der Vekens, N. V. a. n., Beckers, R., Renard, Y., Muysoms, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Descriptive+anatomy+of+the+porcine+ventral+abdominal+wall+as+a+basis+for+training+ventral+hernia+repair+techniques.">Descriptive anatomy of the porcine ventral abdominal wall as a basis for training ventral hernia repair techniques.</a> J Abdom Wall Surg. 3, (2024).</li><li>Luna, S. P. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Validation+of+the+UNESP-Botucatu+pig+composite+acute+pain+scale+(UPAPS).">Validation of the UNESP-Botucatu pig composite acute pain scale (UPAPS).</a> PLoS One. 15 (6), e0233552 (2020).</li><li>Arruda, P. H. E., Gauger, P., Zimmerman, J. J., Karriker, L. A., Ramirez, A., Schwartz, K. J., Stevenson, G. W., Zhang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimizing+sample+selection,+collection,+and+submission+to+optimize+diagnostic+value.">Optimizing sample selection, collection, and submission to optimize diagnostic value.</a> Diseases of swine. 11th ed. , 98-111 (2019).</li><li>Althouse, G. C., Kauffold, J., Rossow, S., Zimmerman, J. J., Karriker, L. A., Ramirez, A., Schwartz, K. J., Stevenson, G. W., Zhang, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Diseases+of+the+reproductive+system.">Diseases of the reproductive system.</a> Diseases of swine. 11th ed. , 373-392 (2019).</li><li>Hao, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Porcine+skin-derived+stem+cells+can+serve+as+donor+cells+for+nuclear+transfer.">Porcine skin-derived stem cells can serve as donor cells for nuclear transfer.</a> Cloning Stem Cells. 11 (1), 101-109 (2010).</li><li>Rim, C. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+the+number+of+transferred+embryos,+the+interval+between+nuclear+transfer+and+embryo+transfer,+and+the+transfer+pattern+on+pig+cloning+efficiency.">The effect of the number of transferred embryos, the interval between nuclear transfer and embryo transfer, and the transfer pattern on pig cloning efficiency.</a> Anim Reprod Sci. 143 (1-4), 91-96 (2013).</li><li>Choi, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Production+of+heterozygous+alpha+1,3-galactosyltransferase+(GGTA1)+knock-out+transgenic+miniature+pigs+expressing+human+CD39.">Production of heterozygous alpha 1,3-galactosyltransferase (GGTA1) knock-out transgenic miniature pigs expressing human CD39.</a> Transgenic Res. 26 (2), 209-224 (2017).</li><li>Shim, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+immune+reactivity+of+GGTA1/CMAH/A3GALT2+triple+knockout+Yucatan+miniature+pigs.">Human immune reactivity of GGTA1/CMAH/A3GALT2 triple knockout Yucatan miniature pigs.</a> Transgenic Res. 30 (5), 619-634 (2021).</li><li>Glanzner, W. G., Rissi, V. B., Bordignon, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Somatic+cell+nuclear+transfer+in+pigs.">Somatic cell nuclear transfer in pigs.</a> Methods Mol Biol. 2647, 197-210 (2023).</li><li>Kim, J. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+production+efficiency+of+cloned+transgenic+Yucatan+miniature+pigs+according+to+recipient+breeds+with+embryo+transfer+conditions.">Analysis of production efficiency of cloned transgenic Yucatan miniature pigs according to recipient breeds with embryo transfer conditions.</a> Theriogenology. 218, 193-199 (2024).</li><li>Navarro-Serna, S., Pi&#241;eiro-Silva, C., Romar, R., Parrington, J., Gadea, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Generation of Gene Edited Pigs. , 71-130 (2022).</li><li>Geisert, R. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+conceptus+elongation+in+the+pig:+An+interleukin+1+beta+2+and+estrogen-regulated+phenomenon.">Rapid conceptus elongation in the pig: An interleukin 1 beta 2 and estrogen-regulated phenomenon.</a> Mol Reprod Dev. 84 (9), 760-774 (2017).</li><li>Besenfelder, U., M&#246;dl, J., M&#252;ller, M., Brem, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endoscopic+embryo+collection+and+embryo+transfer+into+the+oviduct+and+the+uterus+of+pigs.">Endoscopic embryo collection and embryo transfer into the oviduct and the uterus of pigs.</a> Theriogenology. 47 (5), 1051-1060 (1997).</li><li>Hazeleger, W., Kemp, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+developments+in+pig+embryo+transfer.">Recent developments in pig embryo transfer.</a> Theriogenology. 56 (8), 1321-1331 (2001).</li><li>Youngs, C. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+influencing+the+success+of+embryo+transfer+in+the+pig.">Factors influencing the success of embryo transfer in the pig.</a> Theriogenology. 56 (8), 1311-1320 (2001).</li><li>Wieczorek, J., Koseniuk, J., Mandryk, I., Poniedzia&#322;ek-Kempny, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Piglets+born+after+intrauterine+laparoscopic+embryo+transfer.">Piglets born after intrauterine laparoscopic embryo transfer.</a> Pol J Vet Sci. 18 (2), 425-431 (2015).</li><li>Br&#252;ssow, K. P., R&#225;tky, J., Antosik, P., Kempisty, B., Ja&#347;kowski, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Embryo+transfer+in+swine+-+an+indispensable+key+for+the+application+of+reproduction+techniques.">Embryo transfer in swine - an indispensable key for the application of reproduction techniques.</a> Electron J Pol Agric Univ. 21 (3), (2018).</li><li>Cadwallader, J. A., Alley, M. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Malignant+hyperthermia+in+a+crossbred+landrace-large+white+pig.">Malignant hyperthermia in a crossbred landrace-large white pig.</a> N Z Vet J. 23 (9), 207-211 (1975).</li><li>Fatehi Hassanabad, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prevention+of+postoperative+adhesions:+A+comprehensive+review+of+present+and+emerging+strategies.">Prevention of postoperative adhesions: A comprehensive review of present and emerging strategies.</a> Biomolecules. 11 (7), 1027 (2021).</li><li>Shi, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+embryo+handling+and+transfer+method+on+pig+cloning+efficiency.">Influence of embryo handling and transfer method on pig cloning efficiency.</a> Anim Reprod Sci. 154, 121-127 (2015).</li><li>Castagna, C. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ovarian+cysts+and+their+consequences+on+the+reproductive+performance+of+swine+herds.">Ovarian cysts and their consequences on the reproductive performance of swine herds.</a> Anim Reprod Sci. 81 (1-2), 115-123 (2004).</li><li>Knox, R. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Factors+influencing+follicle+development+in+gilts+and+sows+and+management+strategies+used+to+regulate+growth+for+control+of+estrus+and+ovulation.">Factors influencing follicle development in gilts and sows and management strategies used to regulate growth for control of estrus and ovulation.</a> J Anim Sci. 97 (4), 1433-1445 (2019).</li><li>Galvin, J. M., Killian, D. B., Stewart, A. N. V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+procedure+for+successful+nonsurgical+embryo+transfer+in+swine.">A procedure for successful nonsurgical embryo transfer in swine.</a> Theriogenology. 41 (6), 1279-1289 (1994).</li><li>Lins, R. D. A. U., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Use+of+cyanoacrylate+in+the+coaptation+of+edges+of+surgical+wounds.">Use of cyanoacrylate in the coaptation of edges of surgical wounds.</a> An Bras Dermatol. 87 (6), 871-876 (2012).</li><li>Horvath-Pereira, B. d. e. O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Case+report:+An+innovative+non-invasive+technique+to+manage+shell+injuries+in+C.+carbonarius.">Case report: An innovative non-invasive technique to manage shell injuries in C. carbonarius.</a> Front Vet Sci. 9, 930419 (2022).</li></ol>

None of the authors disclose any conflict of interest

The surgical method described has been previously performed by other research groups working with the production of cloned pigs or genetically modified cloned pigs, with reports of births after the implementation of this technique15,16,17,18,19,20. The pregnancy and birth rates of SCNT embryos produced from various cell lines...

Pigs are an excellent experimental model for biomedical research due to their anatomical, physiological, and genetic similarities to humans1. These animals have often been used in research related to xenotransplantation, with the intention of producing organs, cells, or tissues that promote a low risk of rejection when transplanted into humans. Xenotransplantation research aims to increase the organ supply for human transplantation, thus reducing the waiting list of patients2.
The production of pigs for xenotransplantation involves several steps, including the production of clones from genetically edited swine cells. After the in vitro production of genetically modified cloned embryos, the embryos are transferred to the reproductive system of a sow with a synchronized estrous cycle to prepare the uterine physiology for the reception and gestation of the new conceptus3.
Embryo transfer in pigs can be performed by non-invasive or invasive methods4. Among the non-invasive methods is transcervical transfer, which does not require any surgical intervention. However, this method is restricted to transferring embryos at later development stages (i.e., morula or blastocyst stages) and does not allow for the precise determination of the catheter insertion site or embryo deposition5. Laparoscopy and laparotomy are considered invasive methods of embryo transfer. Laparoscopy is less invasive but requires specific and costly equipment, and its efficiency varies considerably (from less than 20% to over 80%) due to various factors such as difficulty in manipulating reproductive structures and the type of catheter used4,6. Therefore, transfer via laparoscopy is still less efficient compared to surgical transfer methods via laparotomy4.
Embryo transfer surgery in sows via laparotomy is a relatively simple and quick procedure, typically taking about 30 min. However, it should be performed in a surgical center equipped with inhalation anesthesia apparatus and a specialized team. For commercial pig strains (such as Landrace, Large White, or their crossbreeds), special equipment like hoists for lifting gilts and a wide, sturdy surgical table are necessary due to the animals&#39; considerable weight (around 130-150 kg).
For the surgery to be successful, the embryos must be evaluated for their stage of development beforehand. Embryos of up to 4 cells are recommended to be transferred into the uterine tube. Embryos at stages beyond 4 cells, such as morulas and blastocysts, should be transferred to the uterine horn7,8.
Although research groups worldwide are performing the production and surgical transfer of genetically modified cloned pig embryos, there are still no well-defined protocols demonstrating this procedure through videos. This approach is crucial for the success of gestation, as the technique requires precise deposition of embryos in the exact location of the oviduct, involving the localization of the tubal ostium and the introduction of the pipette containing the embryos. This technique can be better understood through explanatory videos of the entire procedure. Therefore, this article aims to demonstrate the laparotomy surgery for transferring cloned embryos to the oviduct of gilts, an essential prerequisite for the future production of genetically modified pigs to be used for xenotransplantation or other related purposes.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>0.25 mL straw</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>1 mL syringe</td>
			<td>Descarpack</td>
			<td>341001</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>10 mL syringe</td>
			<td>Descarpack</td>
			<td>324601</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>20 mL syringe</td>
			<td>Descarpack</td>
			<td>324801</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>3 mL syringe</td>
			<td>Descarpack</td>
			<td>324201</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>5 mL syringe</td>
			<td>Descarpack</td>
			<td>324401</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>60 mL syringe</td>
			<td>Descarpack</td>
			<td>323201</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>9 mm endotracheal tube</td>
			<td>Rusch</td>
			<td>112482-000090</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Allis forceps</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Amox LA</td>
			<td>JA Sa&#250;de Animal</td>
			<td>MAPA registration: 8.781/2004</td>
			<td>Pharmaceutical drug</td>
		</tr>
		<tr>
			<td>Bakhaus forceps</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Catheter 20G</td>
			<td>Descarpack</td>
			<td>362401</td>
			<td>Catheter for intravenous access</td>
		</tr>
		<tr>
			<td>Cetamin</td>
			<td>Agener Uni&#227;o</td>
			<td>MAPA registration: SP-000292-5.000011&#160;</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Conductive clinical gel</td>
			<td>RMC</td>
			<td>ANVISA registration: 80122200013</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Dipyrone D-500</td>
			<td>Zoetis</td>
			<td>MAPA registration: SP0000728-46</td>
			<td>Pharmaceutical drug</td>
		</tr>
		<tr>
			<td>Disposable scalpel n. 22</td>
			<td>Wiltex</td>
			<td>ANVISA registration: 10150470565</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Disposable sterile sponge-brush</td>
			<td>Rioquimica</td>
			<td>7.89778E+12</td>
			<td>Surgical asepsis</td>
		</tr>
		<tr>
			<td>Easy-Scan:Go</td>
			<td>IMV</td>
			<td>ESCG01</td>
			<td>Ultrasound</td>
		</tr>
		<tr>
			<td>Endozime AW Plus</td>
			<td>Ruhof</td>
			<td>34514</td>
			<td>Detergent for surgical instruments</td>
		</tr>
		<tr>
			<td>Fentanil (Fentanest)</td>
			<td>Crist&#225;lia</td>
			<td>ANVISA registration: 1029800810159</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Gosset retractor</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Halstead-mosquito hemostatic forceps</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Healing ointment - Unguento Pearson</td>
			<td>Pearson SA</td>
			<td>MAPA registration: SP0000094-16</td>
			<td>Pharmaceutical drug</td>
		</tr>
		<tr>
			<td>hydrogen peroxide solution</td>
			<td>Rioquimica</td>
			<td>ANVISA registration: 218690015</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Isofluorane</td>
			<td>Biochimico</td>
			<td>ANVISA registration: 100630222</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>IV Drip set extensor</td>
			<td>generic</td>
			<td>-</td>
			<td>Fluid therapy</td>
		</tr>
		<tr>
			<td>IV Macro drip set</td>
			<td>Descarpack</td>
			<td>410301</td>
			<td>Fluid therapy</td>
		</tr>
		<tr>
			<td>Lactofur (ceftiofur)</td>
			<td>Ourofino SA</td>
			<td>MAPA registration: SP0000051-50</td>
			<td>Pharmaceutical drug</td>
		</tr>
		<tr>
			<td>Laringoscope</td>
			<td>-</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Maxicam 2%</td>
			<td>Ourofino SA</td>
			<td>MAPA registration: SP0000051-69</td>
			<td>Pharmaceutical drug</td>
		</tr>
		<tr>
			<td>Micropore adhesive 5 cm x 10 cm&#160;</td>
			<td>generic</td>
			<td>1530</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Midazolam</td>
			<td>Hipolabor</td>
			<td>ANVISA registration: 1134301430035</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Multi-Way IV Infusion Set</td>
			<td>Descarpack</td>
			<td>413201</td>
			<td>Fluid therapy</td>
		</tr>
		<tr>
			<td>Needle 40 mm x 1.2 mm</td>
			<td>Descarpack</td>
			<td>353601</td>
			<td>Sterile needle for applying medicines and anesthetics.</td>
		</tr>
		<tr>
			<td>Needle 40 mm x 1.6 mm</td>
			<td>Wiltex</td>
			<td>ANVISA registration: 10150470664</td>
			<td>Sterile needle for applying medicines and anesthetics.</td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Nylon 2 suture trhead</td>
			<td>Shalon Medical</td>
			<td>N502CTI40</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Ordinary pen</td>
			<td>generic</td>
			<td>-</td>
			<td>Regular pen for taking notes</td>
		</tr>
		<tr>
			<td>Physiological solution 0.9% 500 mL bag</td>
			<td>JP Farma</td>
			<td>MS:1.0491.0070</td>
			<td>Fluid therapy</td>
		</tr>
		<tr>
			<td>Polyglycolic acid 2 suture thread</td>
			<td>Atramat</td>
			<td>G4099-75H</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Potassium chloride</td>
			<td>Samtec</td>
			<td>ANVISA registration: 1559200010139</td>
			<td>Parenteral drug</td>
		</tr>
		<tr>
			<td>Procedure gloves</td>
			<td>Descarpack</td>
			<td>122401</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Propofol (Provive)</td>
			<td>Uni&#227;o Qu&#237;mica</td>
			<td>ANVISA registration: 1049714490057</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Ringer lactate solution 500 mL bag</td>
			<td>JP Farma</td>
			<td>MS:1.0491.0061</td>
			<td>Fluid therapy</td>
		</tr>
		<tr>
			<td>Riohex 0.5% clorexidine alcohol solution</td>
			<td>Rioquimica</td>
			<td>218690356</td>
			<td>Surgical asepsis</td>
		</tr>
		<tr>
			<td>Riohex 2% clorexidine solution with surfactant</td>
			<td>Rioquimica</td>
			<td>218690356</td>
			<td>Surgical asepsis</td>
		</tr>
		<tr>
			<td>Scalp 21 G</td>
			<td>Descarpack</td>
			<td>421201</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Shoe covers</td>
			<td>generic</td>
			<td>-</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Sterile compresses</td>
			<td>Cremer</td>
			<td>ANVISA registration: 10071150065</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Sterile gauze pad</td>
			<td>Procitex</td>
			<td>ANVISA registration: 80245210083</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Sterile surgical drapes 140 cm x 90 cm&#160;</td>
			<td>Venkuri</td>
			<td>7010003</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Sterile surgical drapes 150 cm x 190 cm&#160;</td>
			<td>PolarFix</td>
			<td>F00208</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Sterile surgical gloves</td>
			<td>Mucambo</td>
			<td>CA: 39.317</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Stethoscope</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical equipment</td>
		</tr>
		<tr>
			<td>Surgical cap</td>
			<td>Descarpack</td>
			<td>93201</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Surgical gown</td>
			<td>Descarpack</td>
			<td>231101</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Surgical mask</td>
			<td>Descarpack</td>
			<td>110701</td>
			<td>Personal Protective Equipment (PPE)</td>
		</tr>
		<tr>
			<td>Surgical scissors blunt-blunt</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Surgical scissors sharp-sharp</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical instrument</td>
		</tr>
		<tr>
			<td>Surgical stapler</td>
			<td>Tradevet</td>
			<td>-</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Tekbond super glue</td>
			<td>Tek Bond</td>
			<td>78072720030</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Thermometer</td>
			<td>generic</td>
			<td>-</td>
			<td>Surgical equipment</td>
		</tr>
		<tr>
			<td>Three Way Stopcock</td>
			<td>Solidor</td>
			<td>374</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Tramadol hydrocloride</td>
			<td>Agener Uni&#227;o</td>
			<td>MAPA registration: SP-000292-5.000002</td>
			<td>Anesthetic</td>
		</tr>
		<tr>
			<td>Transparent film dressing</td>
			<td>Skinupper</td>
			<td>ANVISA registration: 82307460014</td>
			<td>Surgical material</td>
		</tr>
		<tr>
			<td>Waterproof adhesive 10 cm x 4.5 cm&#160;</td>
			<td>generic</td>
			<td>364828</td>
			<td>Surgical material</td>
		</tr>
	</tbody>
</table>

embryo transfer surgery via laparotomy in gilts

This protocol aims to demonstrate the surgical technique for transferring cloned pig embryos to the oviduct, a method widely used in the production of genetically modified pigs for biomedical research. Nine gilts underwent hormonal synchronization and laparotomy for the transfer of cloned embryos produced by somatic cell nuclear transfer (SCNT) at stages of up to 4 cells on day 2 to the oviduct. Gestational diagnosis was conducted via ultrasound examination 30 days post-transfer surgery. Six out of the nine operated gilts exhibited signs of pregnancy on ultrasound examination. However, as there was no progression in fetal development as assessed by echography, the gilts underwent necropsy at 60 days for the collection of biological material and assessment of the reproductive system. Adhesions were observed in the uterine horns, ovaries, and oviducts. From the uterine lumen of two of the euthanized gilts, one and four embryonic structures with gestational ages ranging between 12 to 20 days were obtained. Despite the absence of live piglets, likely attributed to the low-efficiency rate of transferring cloned pig embryos, which is influenced by various factors, including the number and quality of transferred embryos, the presented surgical technique proved to be rapid and safe.

This article aims to demonstrate the laparotomy surgery for transferring cloned embryos to the oviduct of gilts. All animals remained in an adequate anesthetic plane, without any intraoperative incidents or complications during anesthesia recovery. The gilts took, on average, 2-3 h to stand up after the surgery ended.
All surgical procedures lasted, on average, 44 min. Nine gilts underwent surgery, with an average of 185 cloned embryos transferred per gilt, totaling 1,664 embryos transferred b...

Author Spotlight: Surgical Methods and Outcomes in Oviductal Cloned Pig Embryo Transfers

This protocol describes the application of the surgical technique used to transfer cloned pig embryos via laparotomy in gilts.

Embryo Transfer Surgery via Laparotomy in Gilts

This protocol aims to demonstrate the surgical technique for transferring cloned pig embryos to the oviduct, a method widely used in the production of ...

embryo-transfer-surgery-via-laparotomy-in-gilts

Without Section

Research

JoVE Journal

Medicine

500 Views.  Universidade de São Paulo. This protocol describes the application of the surgical technique used to transfer cloned pig embryos via laparotomy in gilts.

Video: Author Spotlight: Surgical Methods and Outcomes in Oviductal Cloned Pig Embryo Transfers

Single-stage Dynamic Reanimation of the Smile in Irreversible Facial Paralysis by Free Functional Muscle Transfer

Unilateral facial paralysis is a common disease that is associated with significant functional, aesthetic and psychological issues. Though idiopathic facial paralysis (Bell&#8217;s palsy) is the most common diagnosis, patients can also present with a history of physical trauma, infectious disease, tumor, or iatrogenic facial paralysis. Early repair within one year of injury can be achieved by direct nerve repair, cross-face nerve grafting or regional nerve transfer. It is due to muscle atrophy that in long lasting facial paralysis complex reconstructive methods have to be applied. Instead of one single procedure, different surgical approaches have to be considered to alleviate the various components of the paralysis.
The reconstruction of a spontaneous dynamic smile with a symmetric resting tone is a crucial factor to overcome the functional deficits and the social handicap that are associated with facial paralysis. Although numerous surgical techniques have been described, a two-stage approach with an initial cross-facial nerve grafting followed by a free functional muscle transfer is most frequently applied. In selected patients however, a single-stage reconstruction using the motor nerve to the masseter as donor nerve is superior to a two-stage repair. The gracilis muscle is most commonly used for reconstruction, as it presents with a constant anatomy, a simple dissection and minimal donor site morbidity.
Here we demonstrate the pre-operative work-up, the post-operative management, and precisely describe the surgical procedure of single-stage microsurgical reconstruction of the smile by free functional gracilis muscle transfer in a step by step protocol. We further illustrate common pitfalls and provide useful tips which should enable the reader to truly comprehend the procedure. We further discuss indications and limitations of the technique and demonstrate representative results.

The use of the masseteric nerve as donor nerve represents a single-stage alternative to the criterion standard two-stage procedure of cross-facial nerve grafting and free muscle transfer in facial paralysis. We provide a detailed description to safely perform this technique with a gracilis muscle transfer and discuss indications and limitations.

Unilateral facial paralysis is a common disease that is associated with significant functional, aesthetic and psychological issues. Though idiopathic ...

Uncontrolled Hemorrhagic Shock Modeled via Liver Laceration in Mice with Real Time Hemodynamic Monitoring

Uncontrolled hemorrhage is an important cause of preventable deaths among trauma patients. We have developed a murine model of uncontrolled hemorrhage via a liver laceration that results in consistent blood loss, hemodynamic alterations, and survival.
Mice undergo a standardized resection of the left-middle lobe of the liver. They are allowed to bleed without mechanical intervention. Hemostatic agents can be administered as pre-treatment or rescue therapy depending on the interest of the investigator. During the time of hemorrhage, real-time hemodynamic monitoring via a left femoral arterial line is performed. Mice are then sacrificed, blood loss is quantified, blood is collected for further analysis, and organs are harvested for analysis of injury. Experimental design is described to allow for simultaneous testing of multiple animals.
Liver hemorrhage as a model of uncontrolled hemorrhage exists in the literature, primarily in rat and porcine models. Some of these models utilize hemodynamic monitoring or quantify blood loss but lack consistency. The present model incorporates quantification of blood loss, real-time hemodynamic monitoring in a murine model that offers the advantage of using transgenic lines and a high-throughput mechanism to further investigate the pathophysiologic mechanisms in uncontrolled hemorrhage.

Uncontrolled hemorrhage, an important cause of mortality among trauma patients, can be modeled using a standard liver laceration in a murine model. This model results in consistent blood loss, survival, and allows for testing hemostatic agents. This article provides the step-by-step process to perform this valuable model.

Uncontrolled hemorrhage is an important cause of preventable deaths among trauma patients. We have developed a murine model of uncontrolled hemorrhage via ...

Emergency Undocking in Robotic Surgery: A Simulation Curriculum

The following is a training platform to allow robotic surgeons to develop the skills necessary to lead an interprofessional team in emergency undocking of a robotic system. In traditional robotic training for surgeons, a brief web based overview of performing an emergency undocking is provided during initial introductory training to the robotics system. During such a process, there is no training in delineation of interdisciplinary roles for operating room (OR) personnel. The training presented here uses formative simulation and debriefing followed by a lecture. For the simulation, a modified gynecologic simulator is draped in a steep trendelenburg position consistent with most gynecologic laparoscopic surgery. The training torso is modified using tubing hooked to pressure bags of red food colored IV fluid used to simulate a catastrophic vessel injury on demand. Positioned throughout the operating room setting is an interprofessional team consisting of embedded standardized persons (ESPs) to fulfill the roles of a circulating nurse, scrub nurse, anesthesiologist, and bedside assist surgeon. Robotic surgeons are presented a case scenario necessitating emergency undocking, and given control of the robotic instruments. The scenario is terminated following either successful completion of an emergency undock, or at five minutes due to the emergent nature of the case. A debriefing session with hands on training of the steps to emergency undocking, necessary equipment, troubleshooting techniques, and operating room personnel roles follows the simulation. The learners are presented with a short lecture reemphasizing the material presented in the debriefing for their own self-study. This training results in improved time accessing the patient, improved knowledge, confidence, and completion of critical actions, and can be replicated in most institutions. All robotic surgeons should be able to demonstrate competence in this crucial intervention. A limitation of the curriculum is ability to access the in-situ environment for training purposes.

This training platform is designed to allow robotic surgeons to develop the skills necessary to lead an interprofessional team in emergency undocking of the robotic system. Training includes the utilization of the technology and equipment to perform an emergency undocking, as well as delineation of roles for such a scenario.

The following is a training platform to allow robotic surgeons to develop the skills necessary to lead an interprofessional team in emergency undocking of ...

Intraoperative Ultrasound in Spinal Surgery

Since the 1980s, there have been several reports for the use of intraoperative ultrasound as a useful adjunct in spinal surgery. However, with the advent of newer cutting-edge imaging modalities, the use of intraoperative ultrasound in spine surgery has largely fallen out of favor. Despite this, intraoperative ultrasound continues to provide several advantages over other intraoperative techniques such as magnetic resonance imaging and computed tomography including being more cost-effective, efficient, and easy to operate and interpret. Additionally, it remains the only method for the real-time visualization of soft tissue and pathologies. This paper focuses on the advantages of using intraoperative ultrasound, especially in cases of intradural lesions and lesions ventral to the thecal sac when approaching posteriorly.

Here, we present a protocol on the use of intraoperative ultrasound in spinal surgery, particularly in cases of intradural lesions and lesions in the ventral spinal canal when using a posterior approach.

Since the 1980s, there have been several reports for the use of intraoperative ultrasound as a useful adjunct in spinal surgery. However, with the advent ...

Learning Modern Laryngeal Surgery in a Dissection Laboratory

Surgery for laryngeal malignancies requires millimetric accuracy from the different endoscopic and open techniques available. Practice of this surgery is almost completely reserved to a few referral centers that deal with a large proportion of this pathology. Practice on human specimens is not always possible for ethical, economic, or availability reasons. The aim of this study is to provide a reproducible method for the organization of a laryngeal laboratory on ex vivo animal models where it is possible to approach, learn, and refine laryngeal techniques. Porcine and ovine larynges are ideal, affordable, models to simulate laryngeal surgery given their similarity to the human larynx in their anatomical layout and tissue composition. Herein, the surgical steps of transoral laser surgery, open partial horizontal laryngectomy, and total laryngectomy are reported. The merging of endoscopic and exoscopic views guarantees an inside-out perspective, which is vital for the comprehension of the complex laryngeal anatomy. The method was successfully adopted during three sessions of a dissection course &#34;Lary-Gym&#34;. Further perspectives on robotic surgical training are described.

The purpose of this paper is to illustrate how to organize a reproducible laboratory for laryngeal surgery on affordable and closely similar animal laryngeal models in order to improve anatomical and surgical knowledge and skills.

Surgery for laryngeal malignancies requires millimetric accuracy from the different endoscopic and open techniques available. Practice of this surgery is ...

The Microscopic Transcanal Approach in Stapes Surgery Revisited

The microscopic transcanal (aka transmeatal) surgical approach was first described in the 60s, offering a minimally invasive means of reaching the external auditory canal, the middle ear, and epitympanon. Such an approach avoids a retroauricular or endaural skin incision; however, working through a narrow space needs angled microsurgical instruments and specific training in otologic surgery. The transcanal approach restricts the working space; however, it offers a binocular microscopic vision into the middle ear without extended skin incisions and thus, reducing post-operative pain and bleeding. In addition, this minimally invasive approach avoids scar tissue complications, hypoesthesia of the auricle, and potential protrusion of the pinna. Despite its numerous advantages, this method is still not routinely performed by otologic surgeons. Since this minimally invasive technique is more challenging, there is a need for extensive training in order for it to be widely adopted by otologic surgeons. This article provides step-by-step surgical instructions for stapes surgery and reports possible indications, pitfalls, and limitations using this microscopic transcanal technique.

This article describes the microscopic transcanal technique for stapes surgery, providing step-by-step surgical instructions for familiarizing surgeons with this approach.

The microscopic transcanal (aka transmeatal) surgical approach was first described in the 60s, offering a minimally invasive means of reaching the ...

Gasless Endoscopic Thyroidectomy via the Trans-Axillary Approach

For patients with early, low-risk papillary thyroid carcinoma, an increasing number are opting for endoscopic thyroid surgery due to its ability to achieve favorable therapeutic outcomes while maintaining excellent cosmetic results. Among the available endoscopic procedures, the Gasless Endoscopic Thyroidectomy Trans-axillary Approach (GETTA) has gained popularity among surgeons. This is attributed to its straightforward cavity construction, spacious operating area, precise visual field exposure, and manageable learning curve. However, few studies have provided detailed descriptions of the specific surgical steps involved in GETTA. Drawing from a synthesis of existing literature and our own clinical expertise, we present a comprehensive outline of the GETTA procedure. This process can be categorized into five distinct stages: positioning and incision planning; establishment of surgical cavities; identification and protection of the recurrent laryngeal nerve, inferior parathyroid gland, and central neck dissection; localization and preservation of the superior laryngeal nerve, superior parathyroid gland, and dissection of the thyroid's superior pole; transection of the thyroid isthmus followed by en bloc resection of the thyroid gland and central neck lymph nodes. The five-step approach of GETTA is easy to learn and can be adapted for resecting both benign and malignant thyroid and parathyroid diseases.

Gasless Endoscopic Thyroidectomy via the Trans-Axillary Approach

This protocol presents the procedure for conducting thyroidectomy using the gasless endoscopic thyroidectomy trans-axillary approach (GETTA).

For patients with early, low-risk papillary thyroid carcinoma, an increasing number are opting for endoscopic thyroid surgery due to its ability to ...

Liver Transplantation in Mouse and Analyzing Allograft Patency

Successful Orthotopic Liver Transplantation in Mice Utilizing Microcomputed Tomography Angiography

Microcomputed tomography (microCT) angiography is an invaluable resource to researchers. New advances in this technology have allowed for high-quality images to be obtained of micro-vasculature and are high-fidelity tools in the field of organ transplantation. In this model of orthotopic liver transplantation (OLT) in mice, microCT affords the opportunity to evaluate allograft anastomosis in real time and has the added benefit of not having to sacrifice study animals. The choice of contrast, as well as image acquisition settings, create a high-definition image, which gives researchers invaluable information. This allows for evaluation of the technical aspects of the procedure as well as potentially evaluating different therapeutics over an extended duration of time. In this protocol, we detail an OLT model in mice in a stepwise fashion and finally describe a microCT protocol that can give high-quality images, which aid researchers in in-depth analysis of solid organ transplantation. We provide a step-by-step guide for liver transplantation in a mouse, as well as briefly discuss a protocol for evaluating the patency of the graft through microCT angiography.

Author Spotlight: Enhancing Transplantation Research Through MicroCT Angiography in Murine Models 

In this protocol, we discuss the implementation of a model of successful orthotopic liver transplantation (OLT) in mice. Additionally, adjuvants to further analyze allograft patency after successful OLT in a mouse are discussed as well, specifically utilizing microcomputed tomography (microCT) scans.

Microcomputed tomography (microCT) angiography is an invaluable resource to researchers. New advances in this technology have allowed for high-quality ...

Tracheotomy-Based Endotracheal Intubation Protocol for Non-Survival Rat Models

Endotracheal Intubation via Tracheotomy and Subsequent Thoracotomy in Rats for Non-Survival Applications

Endotracheal intubation and subsequent ventilation are often basic requirements for translational research in rat models for various interventions that require controlled or high ventilation pressures or access to the thoracic cavity and organs. Conventional endoorotracheal intubation using the anatomically existing route through the mouth is well suited for survival experiments. However, this procedure poses some challenges, including generally higher levels of the required experience and technical skill, more advanced equipment, and greater time effort with relevant intubation failure rates and complications such as tracheal perforation, temporary systemic hypooxygenation, and relevant aerial leakage.
This manuscript, therefore, presents a detailed step-by-step protocol for endotracheal intubation through tracheotomy in non-survival rat models when guaranteed intubation success, constant oxygenation levels, high ventilation pressures, or open thoracotomy are required.
The protocol emphasizes the importance of meticulous surgical technique to ensure consistent and reliable outcomes, especially for researchers who are inexperienced or lack routine in the technique of endoorotracheal intubation via direct laryngoscopy. This procedure is, therefore, expected to minimize animal suffering and unnecessary animal losses.

Here we introduce a standardized procedure for endotracheal intubation via tracheotomy followed by thoracotomy in rats, aimed at enhancing the precision and reproducibility of non-survival applications requiring invasive ventilation and exposure of thoracic organs in in vivo rat models.

Endotracheal intubation and subsequent ventilation are often basic requirements for translational research in rat models for various interventions that ...

Rat Model for Visceral Organ Malperfusion Studies

Controlled Reversible Visceral Arterial Ischemia, Venous Congestion and Combined Malperfusion via Midline Laparotomy in Rats

Besides sepsis and malignancy, malperfusion is the third leading cause of tissue degradation and a major pathomechanism for various medical and surgical conditions. Despite significant developments such as bypass surgery, endovascular procedures, extracorporeal membrane oxygenation, and artificial blood substitutes, tissue malperfusion, especially of visceral organs, remains a pressing issue in patient care. The demand for further research on biomedical processes and possible interventions is high. Valid biological models are of utmost importance in enabling this kind of research. Due to the multifactorial aspects of tissue perfusion research, which include not only cell biology but also vascular microanatomy and rheology, an appropriate model requires a degree of biological complexity that only an animal model can provide, rendering rodents the obvious model of choice. Tissue malperfusion can be differentiated into three distinct conditions: (1) isolated arterial ischemia, (2) isolated venous congestion, and (3) combined malperfusion. This article presents a detailed step-by-step protocol for the controlled and reversible induction of these three types of visceral malperfusion via midline laparotomy and clamping of the abdominal aorta and caval vein in rats, underscoring the significance of precise surgical methodology to guarantee uniform and dependable results. Prime examples of possible applications of this model include the development and validation of innovative intraoperative imaging modalities, such as Hyperspectral Imaging (HSI), to objectively visualize and differentiate malperfusion of gastrointestinal, gynecological, and urological organs.

Author Spotlight: Advancing Spectral Characterization of Physiological and Malperfused Tissues

This article introduces a standardized procedure for controlled, reversible malperfusion of visceral organs in rat models. The aim is to induce these malperfusion states with a high degree of methodological certainty and control while maintaining technical simplicity and error resilience.

Besides sepsis and malignancy, malperfusion is the third leading cause of tissue degradation and a major pathomechanism for various medical and surgical ...