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Hair Root Staining – What Can Hematoxylin Do for Your Laboratory?

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Event Dates
Event Duration
1 hour

Hair evidence collected as part of a forensic investigation has the potential to provide valuable source information through DNA analysis of its root. In the years prior to 2019, the hair examiners at the North Carolina State Crime Laboratory (NCSCL) noticed that hair roots being sent for DNA analysis were not yielding DNA profiles as expected. Recent advancements in the NCSCL Forensic Biology Section’s detection limits prompted the hair examiners to begin researching whether changes needed to be made to the current hair root removal protocol to increase the likelihood of developing a DNA profile from a hair root.

A validation was completed at the NCSCL for the method of Hematoxylin staining to screen roots in the telogen growth phase for DNA analysis. In this study, over 900 head hairs from approximately 15 living donors were examined for the presence of telogen roots. Those roots were then stained using Hematoxylin and examined for the presence of nuclei. The roots were separated into one of five groups based on the number of nuclei present: Group I (1 to 10 nuclei), Group II (11 to 20 nuclei), Group III (21 to 30 nuclei), Group IV (31 to 40 nuclei), and Group V (41 or greater nuclei). A set of 64 hair roots, including a Negative Control (0 nuclei) and a Positive Control (anagen or catagen growth phase hair roots), were sent for quantitative analysis in the Forensic Biology Section. The quantitative data showed a clear delineation between Groups I and II, where 36% of Group I versus 80% of Group II passed the Forensic Biology Section’s quantification cutoff. All samples in the Negative Control, Group I, and the Positive Control were then amplified along with a representative sample in Groups II through V. After amplification, the delineation between the results of Groups I and II maintained constant with 27% of Group I obtaining DNA profiles versus 89% of Group II obtaining DNA profiles. This showed that the cutoff for the minimum number of nuclei required in order to obtain a potential DNA profile at the NCSCL is eleven or more nuclei.

Hematoxylin staining of telogen roots was successfully implemented into hair casework at the NCSCL in March 2019. Before the development of the Hematoxylin root staining screening method, only hairs with roots in active growth phases (anagen or catagen) or telogen roots with follicular tissue were sent for DNA analysis, resulting in 32% of roots yielding DNA results. Following implementation of Hematoxylin staining the success rate has more than doubled to 69%. This increase in hair root DNA results illustrates the utility of Hematoxylin root staining in improving casework efficiency across multiple laboratory disciplines. Through implementation of this methodology, only hair roots with the best potential to develop a DNA profile are sent for DNA testing, thereby decreasing Forensic Biology Section caseload, cost, and time of analysis. In fact, the implementation of Hematoxylin staining into casework has resulted in a 14% reduction in the number of hair roots forwarded to the Forensic Biology Section for DNA analysis. Further, hair examiners can be assured that hairs not meeting methodology thresholds, but still of evidential value, are preserved for any future examinations.

Detailed Learning Objectives

  1. Attendees will learn the value of hair root staining to visualize nuclei prior to DNA analysis.
  2. Attendees will observe the process of staining hair roots using Hematoxylin.
  3. Attendees will understand how the implementation of Hematoxylin root staining in a forensic laboratory can increase DNA yield and casework efficiency while preserving non-viable roots.

Date Created: July 8, 2022