기증자 운영 및 힌드 - 림브 하베스트 <ol><li> 수술 절차에 대한 microsurgical 악기 살균 집합이 활용됩니다. 외과 수술의 시작하기 전에 변경해야 멸균 장갑을 포함하여 적절한 복장을 착용해야합니다. </li><li> 생쥐가 pentobarbital의 intraperitoneal 주사 (50mg/kg IP)와 anesthetized이며 전체 수술을 통해 안정적인 동물 체온을 용이하게 가열 작업 현장에 배치 부정사. </li><li> 작업의 제기 피부 절개하기 전에 각 모두 세 번 살균이며, 살균제 및 알코올. 밖으로 건조에서 동물 각막을 방지하기 위해 눈이 수술하기 전에 안과 연고 (Vidisic)와 윤활 있습니다. </li><li> 피부 절개는 약 사타구니 인대의 수준에서 오른쪽 사타구니에 걸쳐 이루어집니다. 대퇴 혈관이 양극성 소작과 무딘 절개 무료 해부되며 깊은 지점은 8-0 실크 출혈도 잡았 있습니다. </li><li> 헤파린의 300μl (50 IU)를 방지 응고에 대한 성기 정맥을 통해 주입합니다. </li><li> 대퇴 혈관이 사타구니 인대의 수준에 출혈도 잡았 그리고 묶인에 말초 절단됩니다. </li><li> 뒷다리가로 플러시됩니다 심심풀이 (4 ° C) 오분 이상의 대퇴 동맥을 통해 HTK 솔루션 (5ml). </li><li> 모든 근육 그룹이 급격하게 가위로 나누어져 있으며, osteotomy은 대퇴골의 말초 셋째 수준에서 본 회전 전기와 함께 수행됩니다. </li><li> 수확 사지 그러면 이식까지 보존 및 냉장 창고 얼음처럼 차가운 HTK 솔루션 20mL에 빠져들 수 있습니다. </li></ol> 받는 운영 및 이식 <ol><li> 쥐 수술 분야에 pentobarbital과 배치 부정사의 intraperitoneal 주사와 함께 anesthetized 있습니다. </li><li> 피부 절개는 약 사타구니 인대의 수준에서 오른쪽 사타구니에 걸쳐 이루어집니다. 대퇴 혈관이 양극성 소작과 무딘 절개 무료로 해부하고 있습니다. </li><li> 대퇴 혈관이 가능한 말초로 출혈도 잡았지만, 상복부의 선박 이륙하는 근위 있습니다. 혈관 클램프는 동맥과 정맥에 사타구니 인대의 수준에 배치됩니다. </li><li> 0.010 mm, 커프스 길이 :; 및 정맥 (커프 직경 안쪽 커프스 - 벽 두께 1.151 mm 폴리에틸렌 커프스는 동맥 (0.912 mm, 커프스 - 벽 두께 : 0.010 mm, 커프스 길이 2.0 mm 커프 직경 내부) 세웠어요 : 2.0 mm). 선박 루멘의 가장자리는 다음 선박 팽창시키는와 함께 뽑아이며 대퇴 동맥과 정맥은 즉석 (약 2mm)의 전신을 통해 everted과 원주 8-0 실크 봉합사로 고정됩니다. </li><li> 기증자 갈기갈기를위한 공간을 마련하기 위해받는 사지는 근육 그룹의 날카로운 해부와 제거하고 osteotomy는 대퇴골 중간에서 수행됩니다. </li><li> 기증자 사지는 HTK 솔루션에서 제거하고 이식은 18 게이지 바늘 (길이 : 1.5-2.0 cm)를 사용하여 대퇴골의 osteosynthesis로 시작하는 intramedullary의 막대로. </li><li> 복부 근육 그룹은 밀접하게 4-0 Vicryl 또는 Prolene 중단 봉합과 approximated 있습니다. </li><li> 혈관 anastomoses는받는 사람의 정맥 및 커프 복잡한 통해 기증자의 정맥과 동맥을 당기는에 의해 수행됩니다. 원주 8-0 실크 봉합사는 다시 문합을 확보하기 위해 배치됩니다. </li><li> 정맥 클램프는 동맥 클램프 다음 발표하고, reperfusion는 말초 사지와 도보로 색상의 반환뿐만 아니라 동맥을 통해 타악기 흐름에 의해 시각입니다. </li><li> 나머지 지느러미 근육 그룹은 4-0 Vicryl 또는 Prolene 중단 봉합과 함께 다시 approximated 수 있습니다. </li><li> 피부는 4-0 vicryl 또는 prolene 중단 봉합과 함께 닫힙니다. </li><li> 받는 사람 랫은 가열 패드에 위치 및 복구까지 면밀히 모니터링합니다. </li><li> 받는 동물은 세 posttransplant 일에 대한 예방 항생제 (Cefazolin, 20 MG / 일, IM) 및 Buprenex를 (0.1 MG / kg, SC)받을 수 있습니다. </li></ol>

<ol>
	<li>Shapiro, R. I., Cerra, F. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+model+for+reimplantation+and+transplantation+of+a+complex+organ:+the+rat+hind+limb.">A model for reimplantation and transplantation of a complex organ: the rat hind limb.</a> J Surg Res. 24 (6), 501-506 (1978).</li><li>Petruzzo, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+international+registry+on+hand+and+composite+tissue+transplantation.">The international registry on hand and composite tissue transplantation.</a> Transplantation. 86 (4), 487-492 (2008).</li><li>Schneeberger, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=First+forearm+transplantation:+outcome+at+3+years.">First forearm transplantation: outcome at 3 years.</a> Am J Transplant. 7 (7), 1753-1762 (2007).</li></ol>

관심 없음 충돌 선언하지 않습니다.

실험 연구 수십년은 앞서 및 최근 과거에 성공적인 인간의 손으로 이식 활성화합니다. 1978 년, 샤피로 외. 물론 여러 가지 병적인, 생리 및 면역 측면뿐만 아니라, 복합 조직 이식의 기능 결과를 연구에 적합한 쥐에 orthotopic 뒷다리 이식 모델을 개발했습니다. 우리는 비 봉합 커프 기술을 도입하여, 즉 프로 시저의 가장 어려운 부분 혈관 문합을 수정했습니다. 그것이 높은 생존율 (80-90%)와 프로 시?...

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orthotopic hind-limb transplantation in rats

복합 조직 allotransplantation (CTA)는 이제 손, 팔 또는 숫자의 손실 후에 유효한 치료 옵션을 나타내고 재건 수술의 새로운 치료 법인되고 있습니다. 그러나, 장기적으로 높은 선량 다중 약물 치료는 광범위한 immunosuppressive 응용 프로그램을 halters 심각한 부작용의 위험을 베어링, 그래프트 생존을 보장하기 위해 필요합니다. 이 분야에 더 이상 진행은 기본적인 면역학 및 복합 조직 이식의 국소 빈혈 reperfusion의 부상의 이해에 따라 달라질 수 있습니다. 지금까지 쥐에 orthotopic 뒷다리 이식은 재건 이식 (RT)에 대한 선호하는 설치류 모델을했습니다, 그러나, 그것은 vasculature의 reattachment, 뼈, 근육과 신경에게 대한 특별한 microsurgical 기술을 필요로 매우 까다로운 절차입니다. 우리는 뒷다리 사지 이식을 밀고하는 신속하고 안정적​​인 접근을 제공하는이 모델에 대한 혈관 커프 문합 기술을 도입했습니다. 이 기술은 간소화하고 단축 수술을 성공적으로 높은 생존과 낮은 합병증 속도로 작업을 수행하기 위해 기본적인 microsurgical 경험 외과 수 있습니다. 기술은 물론 면역뿐만 아니라 국소 빈혈 reperfusion의 부상 (이리) 연구에 적합한 것 같습니다.

Watch this Scientific Journal Video about Orthotopic Hind-Limb Transplantation in Rats at JoVE.com

Orthotopic Hind-Limb Transplantation in Rats

여기서 우리는 금 본위 것으로 보인다 orthotopic 쥐 뒷다리 - 사지 이식 절차를 설명<em&gt; 생체내에</em복합 조직 allotransplantation 연구&gt; 모델입니다.

쥐에서 Orthotopic 힌드 - 림브 이식

Composite tissue allotransplantation (CTA) now represents a valid therapeutic option after the loss of a hand, forearm or digits and has become a novel ...

orthotopic-hind-limb-transplantation-in-rats

Research

JoVE Journal

Immunology and Infection

11.6K 조회수.  Innsbruck Medical University. 여기서 우리는 금 본위 것으로 보인다 orthotopic 쥐 뒷다리 - 사지 이식 절차를 설명 생체내에 모델입니다.

비디오: Orthotopic Hind-Limb Transplantation in Rats

In vivo Imaging Method to Distinguish Acute and Chronic Inflammation

Inflammation is a fundamental aspect of many human diseases. In this video report, we demonstrate non-invasive bioluminescence imaging techniques that distinguish acute and chronic inflammation in mouse models. With tissue damage or pathogen invasion, neutrophils are the first line of defense, playing a major role in mediating the acute inflammatory response. As the inflammatory reaction progresses, circulating monocytes gradually migrate into the site of injury and differentiate into mature macrophages, which mediate chronic inflammation and promote tissue repair by removing tissue debris and producing anti-inflammatory cytokines. Intraperitoneal injection of luminol (5-amino-2,3-dihydro-1,4-phthalazinedione, sodium salt) enables detection of acute inflammation largely mediated by tissue-infiltrating neutrophils. Luminol specifically reacts with the superoxide generated within the phagosomes of neutrophils since bioluminescence results from a myeloperoxidase (MPO) mediated reaction. Lucigenin (bis-N-methylacridinium nitrate) also reacts with superoxide in order to generate bioluminescence. However, lucigenin bioluminescence is independent of MPO and it solely relies on phagocyte NADPH oxidase (Phox) in macrophages during chronic inflammation. Together, luminol and lucigenin allow non-invasive visualization and longitudinal assessment of different phagocyte populations across both acute and chronic inflammatory phases. Given the important role of inflammation in a variety of human diseases, we believe this non-invasive imaging method can help investigate the differential roles of neutrophils and macrophages in a variety of pathological conditions.

In vivo Imaging Method to Distinguish Acute and Chronic Inflammation

We describe a non-invasive imaging method for distinguishing inflammatory stages. Systemic delivery of luminol reveals areas of acute inflammation dependent upon MPO activity in neutrophils. In contrast, injection of lucigenin allows for visualization of chronic inflammation dependent upon Phox activity in macrophages.

생체 내 이미징 방법에 급성 및 만성 염증을 구별하는

Inflammation is a fundamental aspect of many human diseases. In this video report, we demonstrate non-invasive bioluminescence imaging techniques that ...

연구

면역학 및 감염병학

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

A mouse model of chronic airway infection is a key asset in cystic fibrosis (CF) research, although there are a number of concerns regarding the model itself. Early phases of inflammation and infection have been widely studied by using the Pseudomonas aeruginosa agar-beads mouse model, while only few reports have focused on the long-term chronic infection in vivo. The main challenge for long term chronic infection remains the low bacterial burden by P. aeruginosa and the low percentage of infected mice weeks after challenge, indicating that bacterial cells are progressively cleared by the host.
This paper presents a method for obtaining efficient long-term chronic infection in mice. This method is based on the embedding of the P. aeruginosa clinical strains in the agar-beads in vitro, followed by intratracheal instillation in C57Bl/6NCrl mice. Bilateral lung infection is associated with several measurable read-outs including weight loss, mortality, chronic infection, and inflammatory response. The P. aeruginosa RP73 clinical strain was preferred over the PAO1 reference laboratory strain since it resulted in a comparatively lower mortality, more severe lesions, and higher chronic infection. P. aeruginosa colonization may persist in the lung for over three months. Murine lung pathology resembles that of CF patients with advanced chronic pulmonary disease.
This murine model most closely mimics the course of the human disease and can be used both for studies on the pathogenesis and for the evaluation of novel therapies.

Long Term Chronic Pseudomonas aeruginosa Airway Infection in Mice

We describe the agar-beads method to establish persistent long-term chronic Pseudomonas aeruginosa airway infection in the mouse model.

장기 만성<em&gt; 녹농균</em마우스의&gt;기도 감염

A mouse model of chronic airway infection is a key asset in cystic fibrosis (CF) research, although there are a number of concerns regarding the model ...

Complete Thymectomy in Adult Rats with Non-invasive Endotracheal Intubation

Thymectomy in neonatal rodents is an established and reliable procedure for immunological studies. However, in adult rats, complications of hemorrhage and pneumothorax from pleural disruption can result in a significant mortality rate. This protocol is a simple method of rat thymectomy that utilizes a mini-sternotomy and endotracheal intubation. Intubation is accomplished with a non-invasive and easily reproducible method and allows for positive pressure ventilation to prevent pneumothorax and a controlled airway that allows sufficient time for careful thymus dissection to minimize pleural disruption. A 1.5 cm sternal incision decreases contact with mediastinal vessels and pleura, while still providing full visualization of the thymus. Following exposure of the mediastinum, the thymus is removed by blunt dissection under magnification. The pleural space is then sealed by suture closure of the pre-tracheal muscles followed by the application of surgical glue. The thorax is then closed by suture closure of the sternum, followed by suture closure of the skin. All thymectomies were complete as evidenced by immunohistochemical (IHC) staining of mediastinal tissue, and absence of na&#239;ve T-cells by flow cytometry, and the procedure had a 96% survival rate. This method is suitable when complete thymectomy with minimal complications is desired for further immunological studies in athymic adult rats.

Rodent thymectomy is a valuable technique in immunological research. Here, a protocol for complete thymectomy in adult rats using a mini-sternotomy along with non-invasive intubation and positive pressure ventilation to minimize perioperative morbidity and mortality is described.

비 침습적 기관 내 삽관 성인 쥐에서 전체 흉선

Thymectomy in neonatal rodents is an established and reliable procedure for immunological studies. However, in adult rats, complications of hemorrhage and ...

Murine Hind Limb Long Bone Dissection and Bone Marrow Isolation

Investigation of the bone and the bone marrow is critical in many research fields including basic bone biology, immunology, hematology, cancer metastasis, biomechanics, and stem cell biology. Despite the importance of the bone in healthy and pathologic states, however, it is a largely under-researched organ due to lack of specialized knowledge of bone dissection and bone marrow isolation. Mice are a common model organism to study effects on bone and bone marrow, necessitating a standardized and efficient method for long bone dissection and bone marrow isolation for processing of large experimental cohorts. We describe a straightforward dissection procedure for the removal of the femur and tibia that is suitable for downstream applications, including but not limited to histomorphologic analysis and strength testing. In addition, we outline a rapid procedure for isolation of bone marrow from the long bones via centrifugation with limited handling time, ideal for cell sorting, primary cell culture, or DNA, RNA, and protein extraction. The protocol is streamlined for rapid processing of samples to limit experimental error, and is standardized to minimize user-to-user variability.

Here we present a protocol for the dissection of hind limb long bones (femurs and tibiae) from the laboratory mouse. We further describe a rapid technique for bone marrow isolation from these bones that utilizes centrifugation for removal of bone marrow from the bone marrow space.

쥐 뒷다리 긴 뼈 해부 및 골수 격리

Investigation of the bone and the bone marrow is critical in many research fields including basic bone biology, immunology, hematology, cancer metastasis, ...

Murine Full-thickness Skin Transplantation

Murine full-thickness skin transplantation is a well-established in vivo model to study alloimmune response and graft rejection. Despite its limited application to humans, skin transplantation in mice has been widely employed for transplantation research. The procedure is easy to learn and perform, and it does not require delicate microsurgical techniques nor extensive training. Moreover, graft rejection in this model occurs in a very reproducible immunological reaction and is easily monitored by direct inspection and palpation. In addition, secondary skin transplantation with donor-matched or third-party skin grafts can be performed on more complex transplant models as an alternative and uncomplicated method to assess donor-specific tolerance. The complications are low and are in general limited to anesthesia overdose or respiratory distress after the procedure. Graft failure, on the other hand, occurs commonly as a result of poor preparation of the graft, incorrect positioning in the graft bed, or inappropriate placement of the bandage. In this article, we present a protocol for full-thickness skin transplantation in mice and describe the important steps necessary for a successful procedure.

Murine full-thickness skin transplantation is a well-established model to study rejection in an alloimmune setting. Here, we provide a tutorial of each step involved in performing a BALB/c--&#62;C57BL/6 full-thickness skin transplant.

쥐의 전체 두께 피부 이식

Murine full-thickness skin transplantation is a well-established in vivo model to study alloimmune response and graft rejection. Despite its limited ...

In Vitro and In Vivo Assessment of T, B and Myeloid Cells Suppressive Activity and Humoral Responses from Transplant Recipients

The main concern in transplantation is to achieve specific tolerance through induction of regulatory cells. The understanding of tolerance mechanisms requires reliable models. Here, we describe models of tolerance to cardiac allograft in rat, induced by blockade of costimulation signals or by upregulation of immunoregulatory molecules through gene transfer. Each of these models allowed in vivo generation of regulatory cells such as regulatory T cells (Tregs), regulatory B cells (Bregs) or regulatory myeloid cells (RegMCs). In this manuscript, we describe two complementary protocols that have been used to identify and define in vitro and in vivo regulatory cell activity to determine their responsibility in tolerance induction and maintenance. First, an in vitro suppressive assay allowed rapid identification of cells with suppressive capacity on effector immune responses in a dose dependent manner, and can be used for further analysis such as cytokine measurement or cytotoxicity. Second, the adoptive transfer of cells from a tolerant treated recipient to a newly irradiated grafted recipient, highlighted the tolerogenic properties of these cells in controlling graft directed immune responses and/or converting new regulatory cells (termed infectious tolerance). These methods are not restricted to cells with known phenotypic markers and can be extended to any cell population. Furthermore, donor directed allospecificity of regulatory cells (an important goal in the field) can be assessed by using third party donor cells or graft either in vitro or in vivo. Finally, to determine the specific tolerogenic capacity of these regulatory cells, we provide protocols to assess the humoral anti-donor antibody responses and the capacity of the recipient to develop humoral responses against new or former known antigens. The models of tolerance described can be used to further characterize regulatory cells, to identify new biomarkers, and immunoregulatory molecules, and are adaptable to other transplantation models or autoimmune diseases in rodent or human.

In Vitro and In Vivo Assessment of T, B and Myeloid Cells Suppressive Activity and Humoral Responses from Transplant Recipients

Here, we present a protocol to induce tolerance in transplantation, and assess in vitro and in vivo the suppressive capacity of distinct cell subsets from the recipient and the immune status of the recipient toward donor or exogenous antigens.

생체 외에서 그리고 Vivo에서 T, B 및 골수성 세포 진압 활동 및 이식 받는 체액 응답의 평가

The main concern in transplantation is to achieve specific tolerance through induction of regulatory cells. The understanding of tolerance mechanisms ...

An Immunological Model for Heterotopic Heart and Cardiac Muscle Cell Transplantation in Rats

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications have been reported since the first description in 1964. After 30 years of performing heterotopic heart transplantation in rats, we have developed a simplified surgical approach, which can be easily taught and performed without further surgical training or background.
After dissection of the ascending aorta and the pulmonary artery and ligation of superior and inferior caval and pulmonary veins, the donor heart is harvested and subsequently perfused with ice-cold saline solution supplemented with heparin. After clamping and incising the recipient abdominal vessels, the donor ascending aorta and pulmonary artery are anastomosed to the recipient abdominal aorta and inferior vena cava, respectively, using continuous running sutures.
Depending on different donor-recipient combinations, this model allows analyses of either acute or chronic rejection of allografts. The immunological significance of this model is further enhanced by a novel approach of in-ear injection of vital cardiac muscle cells and subsequent analysis of draining cervical lymphatic tissue.

We describe a model of heterotopic abdominal heart transplantation in rats, implying modifications of current strategies, which lead to a simplified surgical approach. Additionally, we describe a novel rejection model by in-ear injection of vital cardiac muscle cells, allowing further transplant immunological analyses in rats.

쥐에 있는 이종 심혼 및 심장 근육 세포 이식을 위한 면역학 모형

Heterotopic heart transplantation in rats has been a commonly used model for diverse immunological studies for more than 50 years. Several modifications ...

Optimization of the Cuff Technique for Murine Heart Transplantation

Murine cardiac transplantation has been performed for more than 40 years. With advancements in microsurgery, certain new techniques have been used to improve surgical efficiency. In our lab, we have optimized the cuff technique with two major steps. First, we used the inner tube technique to insert a temporary inner tube into the external jugular vein and carotid artery blood vessel to facilitate eversion of the vessel over the cuff. Second, we performed complete heterotopic cardiac transplantation through the collaboration of two experienced surgeons. These modifications effectively reduced the operation time to 25 minutes, with a success rate of 95%. In this report, we describe these procedures in detail and provide a supplemental video. We believe that this report on the improved cuff technique will offer practical guidance for murine heterotopic heart transplantation and will enhance the utility of this mouse model for basic research.

We introduce an inner tube approach to the cuff technique for mouse cervical heterotopic heart transplantation to help evert the vessel over the cuff. We found that cooperation between two experienced surgeons markedly shortens the operation time.

뮤린 심장 이식을 위한 커프 기술의 최적화

Murine cardiac transplantation has been performed for more than 40 years. With advancements in microsurgery, certain new techniques have been used to ...

Bone Marrow Transplantation Procedures in Mice to Study Clonal Hematopoiesis

Clonal hematopoiesis is a prevalent age-associated condition that results from the accumulation of somatic mutations in hematopoietic stem and progenitor cells (HSPCs). Mutations in driver genes, that confer cellular fitness, can lead to the development of expanding HSPC clones that increasingly give rise to progeny leukocytes harboring the somatic mutation. Because clonal hematopoiesis has been associated with heart disease, stroke, and mortality, the development of experimental systems that model these processes is key to understanding the mechanisms that underly this new risk factor. Bone marrow transplantation procedures involving myeloablative conditioning in mice, such as total-body irradiation (TBI), are commonly employed to study the role of immune cells in cardiovascular diseases. However, simultaneous damage to the bone marrow niche and other sites of interest, such as the heart and brain, is unavoidable with these procedures. Thus, our lab has developed two alternative methods to minimize or avoid possible side effects caused by TBI: 1) bone marrow transplantation with irradiation shielding and 2) adoptive BMT to non-conditioned mice. In shielded organs, the local environment is preserved allowing for the analysis of clonal hematopoiesis while the function of resident immune cells is unperturbed. In contrast, the adoptive BMT to non-conditioned mice has the additional advantage that both the local environments of the organs and the hematopoietic niche are preserved. Here, we compare three different hematopoietic cell reconstitution approaches and discuss their strengths and limitations for studies of clonal hematopoiesis in cardiovascular disease.

We describe three methods of bone marrow transplantation (BMT): BMT with total-body irradiation, BMT with shielded irradiation, and BMT method with no pre-conditioning (adoptive BMT) for the study of clonal hematopoiesis in mouse models.

Clonal hematopoiesis is a prevalent age-associated condition that results from the accumulation of somatic mutations in hematopoietic stem and progenitor ...

Purification and Expansion of Mouse Invariant Natural Killer T Cells for in vitro and in vivo Studies

Invariant Natural Killer T (iNKT) cells are innate-like T Lymphocytes expressing a conserved semi-invariant T cell receptor (TCR) specific for self or microbial lipid antigens presented by the non-polymorphic MHC class I-related molecule CD1d. Preclinical and clinical studies support a role for iNKT cells in cancer, autoimmunity and infectious diseases. iNKT cells are very conserved throughout species and their investigation has been facilitated by mouse models, including CD1d-deficient or iNKT-deficient mice, and the possibility to unequivocally detect them in mice and men with CD1d tetramers or mAbs specific for the semi-invariant TCR. However, iNKT cells are rare and they need to be expanded to reach manageable numbers for any study. Because the generation of primary mouse iNKT cell line in vitro has proven difficult, we have set up a robust protocol to purify and expand splenic iNKT cells from the iV&#945;14-J&#945;18 transgenic mice (iV&#945;14Tg), in which iNKT cells are 30 times more frequent. We show here that primary splenic iV&#945;14Tg iNKT cells can be enriched through an immunomagnetic separation process, yielding about 95-98% pure iNKT cells. The purified iNKT cells are stimulated by anti-CD3/CD28 beads plus IL-2 and IL-7, resulting in 30-fold expansion by day +14 of the culture with 85-99% purity. The expanded iNKT cells can be easily genetically manipulated, providing an invaluable tool to dissect mechanisms of activation and function in vitro and, more importantly, also upon adoptive transfer in vivo.

We describe a rapid and robust protocol to enrich invariant natural killer T (iNKT) cells from mouse spleen and expand them in vitro to suitable numbers for in vitro and in vivo studies.

체외 및 생체 내 연구를 위한 마우스 불변 천연 킬러 T 세포의 정화 및 확장

Invariant Natural Killer T (iNKT) cells are innate-like T Lymphocytes expressing a conserved semi-invariant T cell receptor (TCR) specific for self or ...