Name: the preparation of primary hematopoietic cell cultures from murine bone marrow for electroporation
Uploaded: 2009-01-06T23:59:00.000+00:00
Duration: 8 min 15 s
Description: Watch this Scientific Journal Video about The Preparation of Primary Hematopoietic Cell Cultures From Murine Bone Marrow for Electroporation at JoVE.com

대퇴골과 Tibiae에서 골수 수확 <ol><li> 12 주 된 마우스 (우리가 BALB / C 마우스를 사용하고 있지만 어떤 마우스 변형이 절차를 할 수있는)에 - 절차의 첫 단계는 6의 대퇴골과 tibiae에서 골수를 수확하는 것입니다. 시작하려면, CO 2 흡입 (표시되지 않습니다)로 마우스를 안락사. 각각의 뒷다리에서 수확 대퇴골과 tibiae. 그런 다음, 면도날과 고관절 및 무릎 관절을 제거하는 대퇴골의 각 끝을 잘라 골수를 노출합니다. </li><li> 비슷한 방식으로, 무릎 및 발목 인접 지역을 제거하고 골수를 노출 tibiae의 각 끝을 잘라. </li><li> 26 게이지 바늘로 3 ML의 주사기를 가지고 RPMI 10 % FBS, 페니실린, 스트렙토 마이신과 베타 - 메르 캅 토 에탄올 (BME = 100 음)와 보충으로 채우십시오. 핀셋을 사용하여 RPMI 미디어를 포함하는 페트리 접시 위에 뼈가시오. 뼈의 한쪽 끝을로 주사기 바늘을 삽입하여 골수를 제거하는 플런저를 우울. 주사기 바늘은 잔류 골수를 제거하는 뼈 내부 위아래로 이동할 수 있습니다. </li><li> 문화를 확립을 진행 후 모든 뼈에 대해 절차를 반복합니다. </li></ol> 문화 구축 <ol><li> 골수 문화를 구축하기 위해 세포 현탁액으로 조직을 해산시키기 위해 여러 번 다운 플러시 골수를 피펫합니다. </li><li> 50 ML 원뿔 튜브의 상단에 70 미크론 필터를 삽입하고, 필터에 세포 현탁액을 추가합니다. </li><li> 페트리 접시에게 RPMI 한 시간을 씻어 후 70 미크론 필터로 헹굼 추가합니다. </li><li> 1 ML 주사기 플런저의 고무 엔드를 사용하여 70 미크론 필터 위에 남아있는 골수 조각을 갈아서. </li><li> RPMI과 필터를 한 번 씻으십시오. </li><li> 필터를 세척 후 5 분 1,200 RPM의 세포 현탁액을 원심 분리기. </li><li> PBS에서 resuspending하여 세포 펠렛을 씻으십시오. </li><li> hemocytometer를 사용하여 셀을 계산합니다. 우리는 일반적으로 마우스를 한번의 유골 (두 대퇴골 두 개의 tibiae)에서 ~ 90000000 세포를 구하십시오. 세포를 계산하면 5 분 동안 1,200 rpm으로 그들을 다시 원심 분리기. </li><li> RPMI 및 나누어지는 그 때문에 최종 농도가 ML 당 백만 세포는 충분한 RPMI를 포함하는 조직 문화 flasks에 소량의 세포를 Resuspend. </li><li> 관심의 세포 유형의 개발을 촉진하기 크린 시토킨를 추가합니다. 이 경우 시토킨 인터루킨 - 3 (IL - 3 = 10 NG / ML)은 basophils과 마스트 세포의 발전을 촉진하기 위해 추가됩니다. basophils를 얻기위한 10 일 동안 알을 품다. 돛대 세포의 경우, 5 주 동안 알을 품다. 돛대 세포를 생성할 때, 미디어와 IL - 3는 일주일에 한 번씩 변경해야합니다. </li><li> 여기 마스트 세포가 단지 도금 후 나타나는 방법을 좀 봐입니다. 이 이미지는 10 일 인터루킨 - 3의 추가 후 basophils과 마스트 세포로 마스트 골수 세포의 분화를 보여줍니다. </li><li> 원하는 세포 유형이 취득하면, electroporation 단계로 진행하십시오. </li></ol> Electroporating 셀 <ol><li> electroporation 단계를 시작하려면 문화 플라스크에 세포의 수를 결정합니다. </li><li> 세포는 일반적으로 ML 당 10 만명의 밀도에 electroporated되므로, 원뿔 튜브에 필요한 휴대폰 번호를 전송하고 5 분 1,200 RPM에서 튜브를 원심 분리기. </li><li> centrifuging 후, 미디어를 대기음 1X PBS로 세포를 씻어, 5 분 동안 1,200 rpm으로 다시 원심 분리기. </li><li> PBS를 기음과 ML 당 10,000,000 세포의 세포 현탁액을 만드는 바이오 래드 진 Pulser의 electroporation 버퍼를 추가합니다. </li><li> 세포를 resuspending 후, ML 당 10-20 마이크로 그램의 최종 농도에서 원하는 플라스미드 DNA를 추가합니다. </li><li> 96 - 웰 플레이트 electroporation에 선택의 우물에 세포 현탁액의 나누어지는 150 microliters. </li><li> MXcell 플레이트 챔버의 electroporation 판을 넣고 뚜껑을 닫습니다. </li><li> 돛대 세포의 transfection하기 전에, electroporation 프로토콜은 미리 또는 저장 프로토콜을 사용하지 않는 MXcell로 프로그래밍해야합니다. 최적화 실험을 통해 우리는 마스트 세포의 가장 높은 transfection 효율이 사각형 웨이브 펄스 프로토콜을 사용하여 발생하는 것을 발견했습니다. 우리는 2000uF 1000 ohms에서 300V/20ms, 350/15ms, 350V/20ms 및 350V/10ms 스퀘어 웨이브 펄스를 제공하기 위해 접시에 electroporation 조건을 달라질 수 있습니다. 프로토콜 설정하고 장치에 로드된되면, 세포를 electroporate에 &quot;펄스&quot;를 누르십시오. </li><li> electroporation가 완료되면, 조직 문화 접시에 세포를 전송합니다. 우리는 일반적으로 ML 인터루킨 - 3 당 10 나노 그램 300 microliters RPMI를 (10 % FCS, 페니실린, 스트렙토 마이신과 베타 - 메르 캅 토 에탄올과 보충)가 포함된 48 잘 조직 배양 플레이트에 잘 각 150 microliter의 electroporation 샘플을 전송할 수 있습니다. 전지는 37 하룻밤 incubated 아르 ° C, 그때 표현과 억제를 위해 24 시간 후에 assayed. </li></ol> 트랜sfection 결과 플라스미드 ML 당 20 마이크로 그램 GFP를 사용하여 성공적인 electroporation 후 세포의 형광 현미경 이미지는 동영상에 표시됩니다. 약 30 %의 MXcell의 electroporation 시스템 transfection 효율성을 사용하는 것은 얻을 수 있습니다. 이 시스템은 세포 생존을 유지하면서 당신이 당신의 transfection 효율을 극대화하기 위해 조건을 변경할 수 있습니다.

더 많은 게놈 정보를 얻습니다 siRNA와 같은 새로운 도구를 개발로서, 생리학 관련 세포의 사용은 질병 경로, 단백질 - 단백질 상호 작용 및 신호 전달에 대한 우리의 이해를 심화하기 위해 점점 더 중요 해지고있다. 차 전지는 조직이나 체액에서 직접 얻은하고 체외에서 재배하고 있습니다. 이러한 세포는 외인성 유전 물질의 도입을 포함 방법의 숫자 조작하실 수 있습니다. 그것은 기본 세포에 플라스미드 DNA 또는 siRNA을 소개하는 가장 효과적인 방법은 electroporation 것을 점점 더 분명 해지고 있습니다. Electroporation은 transiently 따라서 외인성 핵산은 세포를 입력할 수 있도록 세포막의 투과성을 증가하기 위해 전기 펄스로 세포를 제공합니다. transfection의이 방법은 다른 세포 유형 / 라인, 따라서, 돛대 세포를 transfect하는 데 사용할 수있는뿐만 아니라 다른 골수 - 유래 세포 사이의 차이에 대한 수용하도록 최적화할 수 있습니다. 또한, 바이러스 - 매개 transfection와는 달리, electroporation은 transfected DNA의 크기에 제한을 부과하거나 광범위한 준비가 필요하지 않습니다. 지질 - 중재 transfection 대​​조적으로, electroporation 덜 독성 수 있으며 transfected 핵산의 endosomal 트래핑 발생하지 않습니다. 바이오 래드는 이러한 세포를 위해 특별히 MXcell의 electroporation 시스템을 개발했습니다. MXcell 여러분 transfection의 효율성과 세포 생존을 최대화하기 위해 조건을 변경할 수 있습니다. transfection 효율을 최적화하고 세포 사망, 전압을 포함하여 electroporation 매개 변수의 다수를 최소화하기 위해, 정전 용량, 저항, 및 펄스 길이는 조정과 평가 수 있습니다.

<table><tbody><tr><td>Gene Pulser MXcell Electroporation System</td><td>Bio-Rad</td><td>165-2670</td><td></td></tr><tr><td>Gene Pulser Electroporation Buffer</td><td>Bio-Rad</td><td>165-2677</td><td></td></tr></tbody></table>

the preparation of primary hematopoietic cell cultures from murine bone marrow for electroporation

그것은 electroporation은 기본 세포에 플라스미드 DNA 또는 siRNA을 소개하는 가장 효과적인 방법임을 더욱 분명 해지고 있습니다. 진 Pulser MXcell의 electroporation 시스템과 진 Pulser의 electroporation 버퍼가 특히 포유 동물 세포와 같은 기본 및 줄기 cells.This 비디오와 같은 어려운 transfect 세포로 핵산을 transfect하기 위해 개발되었습니다 것은 murine 골수에서 기본 조혈 세포 문화를 확립하는 방법을 보여줍니다 다음 MXcell 시스템의 electroporation을 위해 그들을 준비합니다. 우리는 대퇴골과 경골을 분리하여 시작합니다. 대퇴골과 경골 모두에서 골수 그런 다음 수확하고 문화가 설립됩니다. 교양 골수 세포는 다음 transfected 및 분석입니다.

Watch this Scientific Journal Video about The Preparation of Primary Hematopoietic Cell Cultures From Murine Bone Marrow for Electroporation at JoVE.com

The Preparation of Primary Hematopoietic Cell Cultures From Murine Bone Marrow for Electroporation

이 절차는 murine 골수에서 조혈 세포 기본 문화를 확립하고 진 Pulser MXCell의 electroporation 시스템을 사용 transfection하여 다음 방법을 설명합니다.

Electroporation에 대한 Murine 골수에서 조혈 세포 차 문화의 준비

It is becoming increasingly apparent that electroporation is the most effective way to introduce plasmid DNA or siRNA into primary cells. The Gene Pulser ...

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비디오: The Preparation of Primary Hematopoietic Cell Cultures From Murine Bone Marrow for Electroporation

Highly Efficient Gene Disruption of Murine and Human Hematopoietic Progenitor Cells by CRISPR/Cas9

Advances in the hematopoietic stem cell (HSCs) field have been aided by methods to genetically engineer primary progenitor cells as well as animal models. Complete gene ablation in HSCs required the generation of knockout mice from which HSCs could be isolated, and gene ablation in primary human HSCs was not possible. Viral transduction could be used for knock-down approaches, but these suffered from variable efficacy. In general, genetic manipulation of human and mouse hematopoietic cells was hampered by low efficiencies and extensive time and cost commitments. Recently, CRISPR/Cas9 has dramatically expanded the ability to engineer the DNA of mammalian cells. However, the application of CRISPR/Cas9 to hematopoietic cells has been challenging, mainly due to their low transfection efficiencies, the toxicity of plasmid-based approaches and the slow turnaround time of virus-based protocols.
A rapid method to perform CRISPR/Cas9-mediated gene editing in murine and human hematopoietic stem and progenitor cells with knockout efficiencies of up to 90% is provided in this article. This approach utilizes a ribonucleoprotein (RNP) delivery strategy with a streamlined three-day workflow. The use of Cas9-sgRNA RNP allows for a hit-and-run approach, introducing no exogenous DNA sequences in the genome of edited cells and reducing off-target effects. The RNP-based method is fast and straightforward: it does not require cloning of sgRNAs, virus preparation or specific sgRNA chemical modification. With this protocol, scientists should be able to successfully generate knockouts of a gene of interest in primary hematopoietic cells within a week, including downtimes for oligonucleotide synthesis. This approach will allow a much broader group of users to adapt this protocol for their needs.

A protocol for fast CRISPR/Cas9-mediated gene disruption in mouse and human primary hematopoietic cells is described in this article. Cas9-sgRNA ribonucleoproteins are introduced via electroporation with sgRNAs generated through in vitro transcription and commercial Cas9. High editing efficiencies are achieved with limited time and financial cost.

CRISPR/Cas9에 의해 Murine 인간과 조 혈 조상 세포의 고효율 유전자 장애

Advances in the hematopoietic stem cell (HSCs) field have been aided by methods to genetically engineer primary progenitor cells as well as animal models. ...

연구

유전학

Isolation and Intravenous Injection of Murine Bone Marrow Derived Monocytes

As a subtype of leukocytes and progenitors of macrophages, monocytes are involved in many important processes of organisms and are often the subject of various fields in biomedical science. The method described below is a simple and effective way to isolate murine monocytes from heterogeneous bone marrow.
Bone marrow from the femur and tibia of Balb/c mice is harvested by flushing with phosphate buffered saline (PBS). Cell suspension is supplemented with macrophage-colony stimulating factor (M-CSF) and cultured on ultra-low attachment surfaces to avoid adhesion-triggered differentiation of monocytes. The properties and differentiation of monocytes are characterized at various intervals. Fluorescence activated cell sorting (FACS), with markers like CD11b, CD115, and F4/80, is used for phenotyping. At the end of cultivation, the suspension consists of 45%&#177; 12% monocytes. By removing adhesive macrophages, the purity can be raised up to 86%&#177; 6%. After the isolation, monocytes can be utilized in various ways, and one of the most effective and common methods for in vivo delivery is intravenous tail vein injection. 
This technique of isolation and application is important for mouse model studies, especially in the fields of inflammation or immunology. Monocytes can also be used therapeutically in mouse disease models.

Here we present a protocol that generates large amounts of murine monocytes from heterogeneous bone marrow for translational applications. In comparison to others, this new method helps reduce the number of sacrificed animals and lowers costs by avoiding expensive methods such as high gradient magnetic cell separation (MACS).

분리 및 설치류 골수의 정맥 주사 유래 단핵구

As a subtype of leukocytes and progenitors of macrophages, monocytes are involved in many important processes of organisms and are often the subject of ...

면역학 및 감염병학

Using the Gene Pulser MXcell Electroporation System to Transfect Primary Cells with High Efficiency

It is becoming increasingly apparent that electroporation is the most effective way to introduce plasmid DNA or siRNA into primary cells. The Gene Pulser MXcell electroporation system and Gene Pulser electroporation buffer (Bio-Rad) were specifically developed to easily transfect nucleic acids into mammalian cells and difficult-to-transfect cells, such as primary and stem cells. We will demonstrate how to perform a simple experiment to quickly identify the best electroporation conditions. We will demonstrate how to run several samples through a range of electroporation conditions so that an experiment can be conducted at the same time as optimization is performed. We will also show how optimal conditions identified using 96-well electroporation plates can be used with standard electroporation cuvettes, facilitating the switch from electroporation plates to electroporation cuvettes while maintaining the same electroporation efficiency. In the video, we will also discuss some of the key factors that can lead to the success or failure of electroporation experiments.

This procedure shows how to use the Gene Pulser MXcell electroporation system to rapidly and easily identify the best electroporation conditions for mouse embryonic fibroblasts (MEFs) or other primary cells. Considerations for troubleshooting are also discussed in the associated video.

고효율 기본 셀을 Transfect하는 진 Pulser Electroporation MXcell의 시스템을 사용하여

생물학

Phenotypic Analysis and Isolation of Murine Hematopoietic Stem Cells and Lineage-committed Progenitors

The bone marrow is the principal site where HSCs and more mature blood cells lineage progenitors reside and differentiate in an adult organism. HSCs constitute a minute cell population of pluripotent cells capable of generating all blood cell lineages for a life-time1. The molecular dissection of HSCs homeostasis in the bone marrow has important implications in hematopoiesis, oncology and regenerative medicine. We describe the labeling protocol with fluorescent antibodies and the electronic gating procedure in flow cytometry to score hematopoietic progenitor subsets and HSCs distribution in individual mice (Fig. 1). In addition, we describe a method to extensively enrich hematopoietic progenitors as well as long-term (LT) and short term (ST) reconstituting HSCs from pooled bone marrow cell suspensions by magnetic enrichment of cells expressing c-Kit. The resulting cell preparation can be used to sort selected subsets for in vitro and in vivo functional studies (Fig. 2).
Both trabecular osteoblasts2,3 and sinusoidal endothelium4 constitute functional niches supporting HSCs in the bone marrow. Several mechanisms in the osteoblastic niche, including a subset of N-cadherin+ osteoblasts3 and interaction of the receptor tyrosine kinase Tie2 expressed in HSCs with its ligand angiopoietin-15 concur in determining HSCs quiescence. "Hibernation" in the bone marrow is crucial to protect HSCs from replication and eventual exhaustion upon excessive cycling activity6. Exogenous stimuli acting on cells of the innate immune system such as Toll-like receptor ligands7 and interferon-&alpha;6 can also induce proliferation and differentiation of HSCs into lineage committed progenitors. Recently, a population of dormant mouse HSCs within the lin- c-Kit+ Sca-1+ CD150+ CD48- CD34- population has been described8. Sorting of cells based on CD34 expression from the hematopoietic progenitors-enriched cell suspension as described here allows the isolation of both quiescent self-renewing LT-HSCs and ST-HSCs9. A similar procedure based on depletion of lineage positive cells and sorting of LT-HSC with CD48 and Flk2 antibodies has been previously described10. In the present report we provide a protocol for the phenotypic characterization and ex vivo cell cycle analysis of hematopoietic progenitors, which can be useful for monitoring hematopoiesis in different physiological and pathological conditions. Moreover, we describe a FACS sorting procedure for HSCs, which can be used to define factors and mechanisms regulating their self-renewal, expansion and differentiation in cell biology and signal transduction assays as well as for transplantation.

A method to analyse the distribution of bone marrow hematopoietic progenitors in flow cytometry as well as to efficiently isolate highly purified hematopoietic stem cells (HSCs) is described. The isolation procedure is essentially based on magnetic enrichment of c-Kit+ cells and cell sorting to purify HSCs for cellular and molecular studies.

Murine 조혈 줄기 세포와 리니지 - 헌신 Progenitors의 Phenotypic 분석 및 절연

The bone marrow is the principal site where HSCs and more mature blood cells lineage progenitors reside and differentiate in an adult organism. HSCs ...

Lentiviral CRISPR/Cas9-Mediated Genome Editing for the Study of Hematopoietic Cells in Disease Models

Manipulating genes in hematopoietic stem cells using conventional transgenesis approaches can be time-consuming, expensive, and challenging. Benefiting from advances in genome editing technology and lentivirus-mediated transgene delivery systems, an efficient and economical method is described here that establishes mice in which genes are manipulated specifically in hematopoietic stem cells. Lentiviruses are used to transduce Cas9-expressing lineage-negative bone marrow cells with a guide RNA (gRNA) targeting specific genes and a red fluorescence reporter gene (RFP), then these cells are transplanted into lethally-irradiated C57BL/6 mice. Mice transplanted with lentivirus expressing non-targeting gRNA are used as controls. Engraftment of transduced hematopoietic stem cells are evaluated by flow cytometric analysis of RFP-positive leukocytes of peripheral blood. Using this method, ~90% transduction of myeloid cells and ~70% of lymphoid cells at 4 weeks after transplantation can be achieved. Genomic DNA is isolated from RFP-positive blood cells, and portions of the targeted site DNA are amplified by PCR to validate the genome editing. This protocol provides a high-throughput evaluation of hematopoiesis-regulatory genes and can be extended to a variety of mouse disease models with hematopoietic cell involvement.

Described are protocols for the highly efficient genome editing of murine hematopoietic stem and progenitor cells (HSPC) by the CRISPR/Cas9 system to rapidly develop mouse model systems with hematopoietic system-specific gene modifications.

질병 모델에서 조혈 세포의 연구를 위한 렌티바이러스 CRISPR/Cas9 매개 게놈 편집

Manipulating genes in hematopoietic stem cells using conventional transgenesis approaches can be time-consuming, expensive, and challenging. Benefiting ...

Economical and Efficient Protocol for Isolating and Culturing Bone Marrow-derived Dendritic Cells from Mice

The demand for dendritic cells (DCs) is gradually increasing as immunology research advances. However, DCs are rare in all tissues. The traditional method for isolating DCs primarily involves inducing bone marrow (BM) differentiation into DCs by injecting large doses (&#62;10 ng/mL) of granulocyte-macrophage colony-stimulating factor/interleukin-4 (GM-CSF/IL-4), making the procedure complex and expensive. In this protocol, using all BM cells cultured in 10 ng/mL GM-CSF/IL-4 medium, after 3-4 half-culture exchanges, up to 2.7 x 107 CD11c+ cells&#160;(DCs) per mouse (two femurs) were harvested with a purity of 80%-95%. After 10 days in culture, the expression of CD11c, CD80, and MHC II increased, whereas the number of cells decreased. The number of cells peaked after 7 days of culture. Moreover, this method only took 10 min to harvest all bone marrow cells, and a high number of DCs were obtained after 1 week of culture.

Here, we present an economical and efficient method to isolate and generate high-purity bone marrow-derived dendritic cells from mice after 7 days of culture with 10 ng/mL GM-CSF/IL-4.

마우스로부터 골수 유래 수지상 세포를 분리하고 배양하기 위한 경제적이고 효율적인 프로토콜

The demand for dendritic cells (DCs) is gradually increasing as immunology research advances. However, DCs are rare in all tissues. The traditional method ...

흉선과 골수에서 기질 세포 분리 최적화

Stromal Cell Isolation From Hematopoietic Organs

Single-cell sequencing has enabled the mapping of heterogeneous cell populations in the stroma of hematopoietic organs. These methodologies provide a lens through which to study previously unresolved heterogeneity at steady state, as well as changes in cell type representation induced by extrinsic stresses or during aging. Here, we present step-wise protocols for the isolation of high-quality stromal cell populations from murine and human thymus, as well as murine bone and bone marrow. Cells isolated through these protocols are suitable for generating high-quality single-cell multiomics datasets. The impacts of sample digestion, hematopoietic lineage depletion, FACS analysis/sorting, and how these factors influence compatibility with single-cell sequencing are discussed here. With examples of FACS profiles indicating successful and inefficient dissociation and downstream stromal cell yields in post-sequencing analysis, recognizable pointers for users are provided. Considering the specific requirements of stromal cells is crucial for acquiring high-quality and reproducible results that can advance knowledge in the field.

저자 스포트라이트: 단일 세포 염기서열 분석을 위한 기질 세포 분리를 사용한 조혈 연구 발전

Here we present protocols that enable isolation of stromal cells from murine bone, bone marrow, thymus and human thymic tissue compatible with single-cell multiomics.

조혈 장기에서 기질 세포 분리(Stromal Cell Isolation from Hematopoietic Organs)

Single-cell sequencing has enabled the mapping of heterogeneous cell populations in the stroma of hematopoietic organs. These methodologies provide a lens ...

Expression of Fluorescent Fusion Proteins in Murine Bone Marrow-derived Dendritic Cells and Macrophages

Dendritic cells and macrophages are crucial cells that form the first line of defense against pathogens. They also play important roles in the initiation of an adaptive immune response. Experimental work with these cells is rather challenging. Their abundance in organs and tissues is relatively low. As a result, they cannot be isolated in large numbers. They are also difficult to transfect with cDNA constructs. In the murine model, these problems can be partially overcome by in vitro differentiation from bone marrow progenitors in the presence of M-CSF for macrophages or GM-CSF for dendritic cells. In this way, it is possible to obtain large amounts of these cells from very few animals. Moreover, bone marrow progenitors can be transduced with retroviral vectors carrying cDNA constructs during early stages of cultivation prior to their differentiation into bone marrow derived dendritic cells and macrophages. Thus, retroviral transduction followed by differentiation in vitro can be used to express various cDNA constructs in these cells. The ability to express ectopic proteins substantially extends the range of experiments that can be performed on these cells, including live cell imaging of fluorescent proteins, tandem purifications for interactome analyses, structure-function analyses, monitoring of cellular functions with biosensors and many others. In this article, we describe a detailed protocol for retroviral transduction of murine bone marrow derived dendritic cells and macrophages with vectors coding for fluorescently-tagged proteins. On the example of two adaptor proteins, OPAL1 and PSTPIP2, we demonstrate its practical application in flow cytometry and microscopy. We also discuss the advantages and limitations of this approach.

In this article, we provide a detailed protocol for the expression of fluorescent fusion proteins in murine bone marrow derived dendritic cells and macrophages. The method is based on the transduction of bone marrow progenitors with retroviral constructs followed by differentiation into macrophages and dendritic cells in vitro.

Murine 골 수 유래 수지상 세포와 대 식 세포에 형광 성 융해 단백질의 표정

Dendritic cells and macrophages are crucial cells that form the first line of defense against pathogens. They also play important roles in the initiation ...

Electroporation에 대한 Murine 골수에서 조혈 세포 차 문화의 준비

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마우스로부터 골수 유래 수지상 세포를 분리하고 배양하기 위한 경제적이고 효율적인 프로토콜

조혈 장기에서 기질 세포 분리(Stromal Cell Isolation from Hematopoietic Organs)

조혈 장기에서 기질 세포 분리(Stromal Cell Isolation from Hematopoietic Organs)

조혈 장기에서 기질 세포 분리(Stromal Cell Isolation from Hematopoietic Organs)

Murine 골 수 유래 수지상 세포와 대 식 세포에 형광 성 융해 단백질의 표정