Amyloidosis is a heterogeneous group of diseases caused by deposition of misfolded proteins, which usually leads to organ dysfunction. Accurate typing of amyloid deposits is of paramount importance because organ involvements and disease prognosis differ widely among different subtypes, and its treatments are type specific. Correct identification of amyloidogenic protein is crucial to proper treatment. Traditional antibody-based diagnostic methods such as immunohistochemistry and immunofluorescence are helpful in amyloid typing, but limitations of those approaches including antibody availability and serum protein contamination impair sensitivity and specificity of diagnosis. Sometimes misdiagnosis can lead to catastrophic therapeutic outcome. Genetic testing is important to confirm the diagnosis of hereditary amyloidosis. Nowadays proteomic analysis has been used as an advanced strategy for amyloid typing and the gold-standard today is laser microdissection followed by mass spectrometry (LMD/MS), which can identify causal protein without additional clinical information. Furthermore, LMD/MS is performed on formalin-fixed paraffin-embedded (FFPE) specimens, thus large scale retrospective studies based on archival material can be conducted. In recent studies, LMD/MS has been proven superior to traditional methods without the drawbacks mentioned above. This proteomic approach provides guarantee of appropriate clinical management and probability of new insights into the mechanism of amyloidosis.In this article the new advances of studies on subtyping of systemic amyloidosis are reviewed.
Amyloidosis ; classification ; pathology ; Humans ; Immunoglobulin Light-chain Amyloidosis ; Laser Capture Microdissection ; Mass Spectrometry ; Plasma Cells ; Proteomics ; methods

AIMTo investigate the method of detecting gene expression in colon tissue at a single cell level.

METHODSIndividual cell(s) were picked up from colon frozen section using laser microdissection. RNA was extracted, reverse transcribed to complementary DNA (cDNA). cDNA was then analyzed by nested reverse transcription polymerase chain reaction (nested RT-PCR) using two pairs of primers.

RESULTSSingle cell(s) were selectively picked up using an ultraviolet laser micromanipulator. RNA was extracted, reverse transcribed and used for nested RT-PCR. Amplification products of cDNA from down to a single cell could be clearly visualized in the agarose gel.

CONCLUSIONThe combined utilization of laser microdissection and nested RT-PCR provides an opportunity to analyze gene expression at single cell(s) level in colon tissue.

Colon ; cytology ; Gene Expression ; Gene Expression Profiling ; methods ; Humans ; Laser Capture Microdissection ; Reverse Transcriptase Polymerase Chain Reaction ; methods ; Single-Cell Analysis

OBJECTIVE: To assess the application value of laser capture microdissection (LCM) technique for isolating a small number of sperm cells from mixture sample. METHODS: Mixture samples were prepared with sperm cells and vaginal epithelia at different concentrations. Both LCM technique and the differential lysis method were employed to obtain sperm cells from the mixture samples, and DNA was extracted by magnetic beads method. STR genotyping was determined using Identifiler kit. RESULTS: The successful STR genotype rate of sperm cells isolated from mixture samples with LCM technique was 92.86% (13/14). The rate of differential lysis method was 7.14% (1/14). The successful rates between the two methods were statistically different (P < 0.05). CONCLUSION LCM technique can effectively exclude the interference of female cell component and isolate a small number of sperm cells to obtain a single male STR genotyping. LCM technique is obviously better than the differential lysis method.
Cell Separation/methods* ; DNA/isolation & purification* ; DNA Fingerprinting/methods* ; Epithelial Cells ; Female ; Forensic Medicine/methods* ; Genotype ; Humans ; Laser Capture Microdissection/methods* ; Male ; Polymerase Chain Reaction/methods* ; Spermatozoa/cytology* ; Staining and Labeling ; Tandem Repeat Sequences ; Vagina/cytology*

This paper reviews the advances of DNA detection on three types of difficult biological specimens including degraded samples, trace evidences and mixed samples. The source of different samples, processing methods and announcements were analyzed. New methods such as mitochondrial test system, changing the original experimental conditions, low-volume PCR amplification and new technologies such as whole genome amplification techniques, laser capture micro-dissection, and mini-STR technology in recent years are introduced.
Biomarkers ; Body Fluids/chemistry* ; DNA/genetics* ; DNA Fingerprinting/methods* ; DNA, Mitochondrial/genetics* ; Forensic Medicine/methods* ; Genome, Human ; Humans ; Laser Capture Microdissection/methods* ; Microsatellite Repeats ; Nucleic Acid Amplification Techniques/methods* ; Polymerase Chain Reaction/methods* ; Reproducibility of Results ; Sample Size