siRNA / miRNA gene silencing Human HNSCC cell line Eph receptor B4

TUNEL assay is the cell death detection method where the biochemical marker of apoptosis is DNA fragmentation. The assay involves the microscopical detection of generated DNA fragments with free 3'-hydroxyl residues. in apoptotic cells using enzyme terminal deoxynucleotidyl transferase (TdT) which adds biotinylated nucleotides at the site of DNA breaks. Major challenges of this method involve proper access of the enzyme which could be hampered by poor permeabilization and/or excessive fixation with cross-linking fixative (common with archival tissue). This issue can be resolved by optimizing the incubation time with Proteinase K or CytoninTM.

Get tips on using Purified Mouse Anti-Human MLH1 Clone G168-728 (RUO) to perform Immunohistochemistry Human - MLH1

Get tips on using Monoclonal Mouse Anti-Human Cytokeratin 20 (Concentrate) Clone Ks20.8 to perform Immunohistochemistry Human - CK20

Get tips on using Human Sonic Hedgehog/Shh N-Terminus Quantikine ELISA Kit to perform ELISA Human - ShhN

Get tips on using Human Retinol binding protein 4 ELISA Kit (RBP4) (ab108897) to perform ELISA Human - RBP4

Get tips on using Human NRG1-beta 1/HRG1-beta 1 DuoSet ELISA to perform ELISA Human - NRG1

Get tips on using Human FABP2 / Fatty Acid-Binding Protein, Intestinal ELISA Kit to perform ELISA Human - FABP2

Get tips on using Human Breast Cancer Susceptibility Protein 2 (BRCA2) ELISA Kit to perform ELISA Human - BRCA2

The formation of DNA from an RNA template using reverse transcription leads to the formation of double-stranded complementary DNA or cDNA. The challenges with this process include 1. Maintaining the integrity of RNA, 2. Hairpin loops or other secondary structures formed by single-stranded RNA can also affect cDNA synthesis, and 3. DNA-RNA hybrids, which may result when the first strand of cDNA is formed. For the first challenge, using workflows that involve proper isolation and storage of RNA, and maintaining a nuclease-free environment helps obtain RNA with ideal 260/230 ratios. Using a reverse transcriptase that can tolerate high temperatures (50-55oC), overcomes obstacles imposed by secondary RNA structures. Finally, RNase H has the ability to hydrolyze RNA before the formation of a second cDNA strand. It is important to ensure that RNase H activity is optimal because higher RNase H activity leads to premature degradation of the RNA template. Many reverse transcriptases offer built-in RNase H activity.

Get tips on using MEGM-Mammary-Epithelial-Cell-Growth-Medium to perform Mammalian cell culture media MCF-10A