What causes false-negative ER and PgR results in breast tumors and how can they be avoided?

Updated: Mar 20, 2019
  • Author: Oudai Hassan, MD; Chief Editor: Chandandeep Nagi, MD  more...
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Quality concerns [6]

The most important concern in testing for ER is false-negative results. These can occur from fixation times that are too short (< 6 h) [9] or too long (weeks), [10] decalcification and mercury-containing fixatives (B5), storage of unstained sections for more than 1 week, and insufficient antigen retrieval.

Insufficient antigen retrieval is thought to be one of the most important factors leading to false-negative findings. [11] Table 4 in the next section ("HER2/neu Status") provides a number of practical tips to avoid false-negative results.

Some of the challenges that may lead to a false-positive result are entrapped normal cells, cytoplasmic positivity, and mistaking the control sample on the same slide with carcinoma.

Avoiding false-negative ER and PgR immunohistochemistry results

Considerations concerning preanalytic/analytic variables are as follows:

  • Appropriate fixation: The time from tissue acquisition to fixation should be as short as possible; specimens should be fixed in neutral buffered formalin for 6-72 hours, sliced at 5- to 10-mm intervals, and placed in a sufficient volume of neutral buffered formalin; the time to fixation and the duration of fixation should be recorded for each sample

  • Appropriate antigen retrieval (to be determined by individual laboratories; at least 25 minutes has been suggested)

  • Appropriate controls (ideally, controls with both high and low levels of ER should be used)

  • Other preanalytic factors that might lead to a false-negative results

Considerations during pathologist interpretation are as follows:

  • When possible, the test should be performed on a block with an internal control, because this internal control will be an indicator of fixation issues; normal benign ducts and lobules show patchy to diffuse ER immunopositivity.

  • ER positivity is expected in virtually all grade 1 tumors, in pure tubular carcinomas, in colloid carcinomas, and in classic lobular carcinoma. [12] Negative results in these tumors should be questioned with repeat testing, if indicated; ER-negative, PgR-positive tumors are very uncommon (< 5%); the test should be repeated, if indicated.

  • If the results on core biopsy are questioned, repeat testing on an excision is warranted.

  • Negative results on unstained sections stored for prolonged periods should be questioned; unstained sections can be stored in a freezer to slow down oxidation-mediated reduction of antigenicity.

  • Negative results in bone biopsies should be questioned because of the possible effects of decalcification or B5 fixative. (Fine-needle aspiration or touch preparation samples are a good alternative.)

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