Navigation Links
Working With Laser Capture Microdissection Samples


Laser Capture Microdissection (LCM) is a method for obtaining pure populations of cells from heterogeneous samples. LCM permits the selection and capture of cells, cell aggregates, and discrete morphological structures from thin tissue sections. For LCM, frozen and OCT embedded, or formalin-fixed and paraffin embedded tissue sections are processed, stained, and dehydrated, then scanned under an inverted microscope. Cells are visualized through a thermoplastic film which is attached to the bottom of an optically clear microfuge tube cap. A laser pulse, directed onto the target cells through the film, melts the film and allows it to flow onto the targeted area where it cools and bonds with the underlying cell(s). The film including the adhered cells or clusters is then lifted. Captured cells can then be used for nucleic acid studies, including SNP analysis, endpoint and real-time RT-PCR, and mRNA expression profiling.


Formalin vs. Frozen Sections
LCM is usually carried out on frozen sections when samples will be used for RNA isolation. This is because formalin fixation introduces crosslinks between nucleic acids and proteins, resulting in fragmentation of the RNA. Freezing avoids crosslinking but introduces a different problem. Endogenous nucleases are not irreversibly inactivated, and can become active during the tissue processing steps (fixation, staining, destaining, and dehydration) carried out after sectioning.


An Improved Protocol for Staining
Ambion scientists performed a detailed analysi s of RNA recovered from LCM sections after various steps of the conventional post-sectioning processing protocols for frozen samples. These studies revealed that significant RNA degradation occurred during exposure of the section to aqueous solutions, presumably due to reactivation of endogenous nucleases. To circumvent this problem, Ambion developed a staining procedure that avoids exposing the sections to water at any step. (See our LCM Staining Kit.) Using the optimized "water-free" tissue processing protocol in conjunction with the RNAqueous-Micro Kit, total RNA of excellent quality, as assessed by microfluidic capillary electrophoresis and real-time RT-PCR, can be obtained from pure populations of cells selected by LCM (Figure 1).

Figure 1. Real-Time RT-PCR of ICM Samples. Mouse spleen (A) and liver (B) samples (1 mg, 3 mg and 10 mg) were dissected by LCM and RNA was isolated using the RNAqueous-Micro Kit according to protocol. The RNA was then used as a template for the amplification of GAPDH by real-time RT-PCR.


back to top


Ordering Information
Cat# Product Name Size 1931 RNAqueous-Micro Kit 50 purifications 1935 LCM Staining Kit Reagents for processing 80 slides
'"/>

Source:


Page: All 1 2 3

Related biology technology :

1. Maintaining High Sensitivity when Working with Small-Volume Samples
2. Isolate and Analyze Total RNA from Cells Harvested by Laser Capture Microdissection
3. Laser microdissection and nucleic acid purification - a Leica - QIAGEN cooperation
4. Laser Capture Microdissection (LCM) for the Analysis of Macrophage Gene Expression from Atherosclerotic Lesions
5. Laser Capture Microdissection (LCM) and Expression Profiling of Long-Range Projection Neurons
6. Differential Gene Expression Analysis of Pure Cell Populations from Frozen UterineTissue Using Laser Capture Microdissection (LCM) and cDNA Microarrays
7. Laser Capture Microdissection of muscle fiber populations and expression analysis by RT-PCR
8. Laser Capture Microdissection of Cells Labeled with Enhanced Green Fluorescent Protein
9. Quantitation of the Protein MIA as a Marker for Chondrocytes in Research Samples
10. Isolate High Quality Total RNA from LCM Samples Suitable for Microarray and qRT-PCR Analysis
11. Preserve Samples for RNA Expression Microarrays RNAlater Around the World
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:12/2/2016)... --  SurePure, Inc. (OTCQB: SURP) a global leader ... concluded an agreement with Tamarack Biotics under which Tamarack ... acquire units of the Company,s patented photopurification technology with ... Concurrently with the option, SurePure has entered ... seek regulatory approvals in the United States ...
(Date:11/30/2016)... 30, 2016 The global Pyrogen ... hold a dominant share in the overall market. The ... Inc., and Merck KGaA, held a lion,s share of ... Research observes that these companies are expected to retain ... that are do not require rabbit pyrogen testing along ...
(Date:11/30/2016)... , Nov. 30, 2016  Tempus, a ... cancer care, and Penn,s Abramson Cancer Center have ... a positive response to immunotherapy treatment based on ... As part of a research collaboration, Tempus ... and melanoma cancer patient data to Penn. Utilizing ...
(Date:11/30/2016)... IN (PRWEB) , ... November 30, 2016 , ... SSCI, ... experts to discuss the implications of the latest FDA guidance on pharmaceutical cocrystals ... 8, 2016 in Cambridge, MA. , The event follows the successful November ...
Breaking Biology Technology:
(Date:6/20/2016)... 2016 Securus Technologies, a leading provider ... public safety, investigation, corrections and monitoring announced that ... has secured the final acceptance by all three ... Access Systems (MAS) installed. Furthermore, Securus will have ... installed by October, 2016. MAS distinguishes between legitimate ...
(Date:6/15/2016)... June 15, 2016 Transparency ... titled "Gesture Recognition Market by Application Market - Global Industry Analysis ... 2024". According to the report, the  global gesture recognition ... 2015 and is estimated to grow at a ... by 2024.  Increasing application of gesture ...
(Date:6/9/2016)... TURKU, Finland , June 9, 2016 ... French National Police deploy Teleste,s video security solution to ensure ... France during the major tournament ... and data communications systems and services, announced today that its ... Police Prefecture to back up public safety across ...
Breaking Biology News(10 mins):