Array Advice - General

View solutions to common problems for arrays:

Solutions to common problems

 Problem 1: Low/no signal on array

Cause:

  1. Poor DNA labelling
  2. Wrong side of the slide was hybridised
  3. Wrong side of the slide was scanned

Solution:

  1. Check labelling efficiency on a Nanodrop
  2. Ensure correct side of slide was hybridised
  3. Check that the correct side of the slide was scanned

 

Problem 2 – Poor signal intensity

Fig 1. Example of poor signal intensityFigure 1. Example of poor signal intensity

Possible causes:

  1. Poor DNA quality or low input
  2. Errors during the labelling reaction e.g. pipetting, purification step
  3. Wash or hybridisation conditions too stringent
  4. Cy5-labeled targets were exposed to light

Solutions:

  1. Re-quantify or purify gDNA.
  2. Ensure protocol is followed.
  3. Hybridisation oven temp should not be greater than 67˚C and slides should be in wash buffer 2 for no more than 1 minute.
  4. Cover tubes with aluminium foil or use amber microcentrifuge tubes.

 

Problem 3 – Poor SNP data

Fig. 2. Examples of good and poor SNP dataFigure 2. Examples of good and poor SNP data

Possible cause: Hybridisation oven/wash buffer 2 temperature incorrect

Solution:

  1. Ensure oven temp is between 65-67˚C by using a calibrated thermometer that is attached to the rotator to mimic temp of the array
  2. Buffer 2 taken out too early from 37˚C incubator
  3. Ensure that the wash area and the 37˚C incubator are close to each other so that the wash buffer 2 temp does not drop too low during transport.

 

CytoSure™ products are for research use only; not for use in diagnostic procedures