Scientific and Regulatory Policy Committee Points to Consider*: Sample Selection,Assay Design,Data Generation and Interpretation,and Reporting Practices for Chromogenic Immunohistochemical (IHC) Assays in Nonclinical Drug Development
Restricted accessResearch articleFirst published online April, 2026
Scientific and Regulatory Policy Committee Points to Consider*: Sample Selection,Assay Design,Data Generation and Interpretation,and Reporting Practices for Chromogenic Immunohistochemical (IHC) Assays in Nonclinical Drug Development
Chromogenic immunohistochemistry (IHC) is an important molecular localization assay in biomedical research and nonclinical drug development, enabling the visualization of specific epitopes within tissues. The methodology is widely used in drug target selection, risk assessment, understanding disease biology, and characterizing histopathological findings in nonclinical studies. The Scientific and Regulatory Policy Committee of the Society of Toxicologic Pathology formed a working group to compile essential information on chromogenic IHC assays not performed in compliance with Good Laboratory Practice (GLP) from nonclinical studies, using relevant literature and the Working Group members’ collective expertise. In this “Points to Consider” article, emphasis is placed on factors influencing IHC data quality, including sample selection, general assay considerations, data generation and interpretation, and effective reporting. The Working Group members deliberated extensively on pertinent topics, aiming to provide specific and practical guidance for pathologists, histologists, and allied scientists engaged in chromogenic IHC assays. While refraining from an exhaustive exploration of the intricate technical details associated with chromogenic IHC, this article offers insights to enhance the accuracy, credibility, and reproducibility of chromogenic IHC, thereby facilitating informed decision-making in the nonclinical development of biomedical products.
AeffnerFWilsonKMartinNT, et al. The “gold standard” paradox in digital image analysis: manual vs. Automated scoring as ground truth. Arch Pathol Lab Med. 2017;141(9):1267-1275. doi:10.5858/arpa.2016-0386-RA.
2.
AeffnerFZarellaMDBuchbinderN, et al. Introduction to digital image analysis in whole-slide imaging: a white paper from the Digital Pathology Association. J Pathol Inform. 2019;10:9. doi:10.4103/jpi.jpi_82_18.
3.
AllredDCHarveyJMBerardoMClarkGM. Prognostic and predictive factors in breast cancer by immunohistochemical analysis. Mod Pathol. 1998;11(2):155-168.
4.
AmgadMStovgaardESBalslevE, et al. Report on computational assessment of tumor infiltrating lymphocytes from the International Immuno-Oncology Biomarker Working Group. NPJ Breast Cancer. 2020;6:16. doi:10.1038/s41523-020-0154-2.
BabaTIshizuAIwasakiS, et al. CD4+/CD8+ macrophages infiltrating at inflammatory sites: a population of monocytes/macrophages with a cytotoxic phenotype. Blood. 2006;107(5):2004-2012. doi:10.1182/blood-2005-06-2345.
7.
BakerM. Reproducibility crisis: blame it on the antibodies. Nature. 2015;521(7552):274-276. doi:10.1038/521274a.
8.
BenczeJSzarkaMKótiB, et al. Comparison of semi-quantitative scoring and artificial intelligence aided digital image analysis of chromogenic immunohistochemistry. Biomolecules. 2021;12(1):19. doi:10.3390/biom12010019.
9.
BeraKSchalperKARimmDLVelchetiVMadabhushiA. Artificial intelligence in digital pathology—new tools for diagnosis and precision oncology. Nat Rev Clin Oncol. 2019;16(11):703-715. doi:10.1038/s41571-019-0252-y.
10.
BlaschkeRJHahurijNDKuijperS, et al. Targeted mutation reveals essential functions of the homeodomain transcription factor Shox2 in sinoatrial and pacemaking development. Circulation. 2007;115(14):1830-1838. doi:10.1161/CIRCULATIONAHA.106.637819.
11.
BledsoeMJGrizzleWE. Use of human specimens in research: the evolving United States regulatory, policy, and scientific landscape. Diagn Histopathol (Oxf). 2013;19(9):322-330. doi:10.1016/j.mpdhp.2013.06.015.
12.
BolonBAeffnerF. A Primer for Oncoimmunology (Immunooncology). Toxicologic Pathology. 2017;45(5):584-588. doi:10.1177/0192623317713318
13.
BolonBFranckeSCaverly RaeJM, et al. Scientific and Regulatory Policy Committee best practices: recommended (“best”) practices for informed (non-blinded) versus masked (blinded) microscopic evaluation in animal toxicity studies. Toxicol Pathol. 2022;50(8):930-941. doi:10.1177/01926233221135563.
14.
BordeauxJWelshAAgarwalS, et al. Antibody validation. Biotechniques. 2010;48(3):197-209. doi:10.2144/000113382.
15.
BressenotAMarchalSBezdetnayaLGarrierJGuilleminFPlénatF. Assessment of apoptosis by immunohistochemistry to active caspase-3, active caspase-7, or cleaved PARP in monolayer cells and spheroid and subcutaneous xenografts of human carcinoma. J Histochem Cytochem. 2009;57(4):289-300. doi:10.1369/jhc.2008.952044.
16.
BuchwalowIBockerW. Antigen detection on tissues using primary antibody raised in the same species. In: BuchwallowIBöckerW, eds. Immunohistochemistry: Basics and Methods. Berlin, Germany: Springer Verlag; 2010:77-81.
17.
BuchwalowISamoilovaVBoeckerWTiemannM. Non-specific binding of antibodies in immunohistochemistry: fallacies and facts. Sci Rep. 2011;1:28. doi:10.1038/srep00028.
18.
BuchwalowIBBöckerW. Immunohistochemistry: Basics and Methods. Berlin, Germany: Springer-Verlag; 2010.
19.
BussiereJLLeachMWPriceKDMounhoBJLightfoot-DunnR. Survey results on the use of the tissue cross-reactivity immunohistochemistry assay. Regul Toxicol Pharmacol. 2011;59(3):493-502. doi:10.1016/j.yrtph.2010.09.017.
20.
CallisGSterchiD. Decalcification of bone: literature review and practical study of various decalcifying agents. Methods, and their effects on bone histology. J Histotechnol. 1998;21(1):49-58. doi:10.1179/his.1998.21.1.49.
21.
Centre for Evidence-Based Medicine, Oxford University. Catalogue of Bias. Accessed November 30, 2025. https://catalogofbias.org/.
22.
ChandPGargASinglaVRaniN. Evaluation of immunohistochemical profile of breast cancer for prognostics and therapeutic use. Niger J Surg. 2018;24(2):100-106.
23.
ChidlowGDaymonMWoodJPCassonRJ. Localization of a wide-ranging panel of antigens in the rat retina by immunohistochemistry: comparison of Davidson’s solution and formalin as fixatives. J Histochem Cytochem. 2011;59(10):884-898. doi:10.1369/0022155411418115.
24.
ChiribogaLCallisGMWangYChlipalaE. Guide for collecting and reporting metadata on protocol variables and parameters from slide-based histotechnology assays to enhance reproducibility. J Histotechnol. 2022;45(4):132-147. doi:10.1080/01478885.2022.2134022.
25.
ChlipalaEABendzinskiCMDornerC, et al. An image analysis solution for quantification and determination of immunohistochemistry staining reproducibility. Appl Immunohistochem Mol Morphol. 2020;28(6):428-436. doi:10.1097/PAI.0000000000000776.
26.
ChlipalaEAButtersMBrousM, et al. Impact of preanalytical factors during histology processing on section suitability for digital image analysis. Toxicol Pathol. 2021;49(4):755-772. doi:10.1177/0192623320970534.
27.
Clinical and Laboratory Standards Institute. Design principles for processing and immunostaining steps. In: Quality Assurance for Design Control and Implementation of Immunohistochemistry Assays; Approved Guideline (CLSI Document I/LA28-A2). 2nd ed.Clinical and Laboratory Standards Institute; 2011:55-80.
28.
CohnM. Degeneracy, mimicry and crossreactivity in immune recognition. Mol Immunol. 2005;42(5):651-655. doi:10.1016/j.molimm.2004.09.010.
29.
ComptonCCRobbJAAndersonMW, et al. Preanalytics and precision pathology: pathology practices to ensure molecular integrity of cancer patient biospecimens for precision medicine. Arch Pathol Lab Med. 2019;143(11):1346-1363. doi:10.5858/arpa.2019-0009-SA.
30.
CoppolaLCianfloneAGrimaldiAM, et al. Biobanking in health care: evolution and future directions. J Transl Med. 2019;17(1):172. doi:10.1186/s12967-019-1922-3.
31.
CrissmanJWGoodmanDGHildebrandtPK, et al. Best practices guideline: toxicologic histopathology. Toxicol Pathol. 2004;31(1):126-131. doi:10.1080/01926230490268756.
32.
DabbsDJ. Diagnostic Immunohistochemistry: Theranostic and Genomic Applications. 6th ed.Amsterdam, The Netherlands: Elsevier; 2021.
33.
DachetFBrownJBValyi-NagyT, et al. Selective time-dependent changes in activity and cell-specific gene expression in human postmortem brain. Sci Rep. 2021;11(1):6078. doi:10.1038/s41598-021-85801-6.
34.
DaiJChenYDaiR, et al. Agonal factors distort gene-expression patterns in human postmortem brains. Front Neurosci. 2021;15:614142. doi:10.3389/fnins.2021.614142.
35.
DelpireE. Research antibodies: do not use them to stain your reputation. Am J Physiol Cell Physiol. 2015;309(11):C707-C708. doi:10.1152/ajpcell.zh0-7843-editorial.2015.
36.
DunstanRWWhartonKAJrQuigleyCLoweA. The use of immunohistochemistry for biomarker assessment—can it compete with other technologies?Toxicol Pathol. 2011;39(6):988-1002. doi:10.1177/0192623311419163.
EconomouMSchöniLHammerCGalvánJAMuellerD-EZlobecI. Proper paraffin slide storage is crucial for translational research projects involving immunohistochemistry stains. Clin Transl Med. 2014;3(1):4. doi:10.1186/2001-1326-3-4.
39.
EliasJM. Immunohistochemical methods. In: Immunohistopathology: A Practical Approach to Diagnosis. 2nd ed.Chicago, IL: ASCP Press; 2003:1-110.
40.
EngelKBMooreHM. Effects of preanalytical variables on the detection of proteins by immunohistochemistry in formalin-fixed, paraffin-embedded tissue. Arch Pathol Lab Med. 2011;135(5):537-543. doi:10.5858/2010-0702-RAIR.1.
41.
Esteva-SociasMArtigaM-JBahamondeO, et al. In search of an evidence-based strategy for quality assessment of human tissue samples: report of the tissue Biospecimen Research Working Group of the Spanish Biobank Network. J Transl Med. 2019;17(1):370. doi:10.1186/s12967-019-2124-8.
42.
FedchenkoNReifenrathJ. Different approaches for interpretation and reporting of immunohistochemistry analysis results in the bone tissue–a review. Diagn Pathol. 2014;9:221. doi:10.1186/s13000-014-0221-9.
43.
FestingMFW. The “completely randomized” and the “randomised block” are the only experimental designs suitable for widespread use in pre-clinical research. Sci Rep. 2020;10(1):17577. doi:10.1038/s41598-020-74538-3.
44.
FitzgibbonsPLBradleyLAFathereeLA, et al. Principles of analytic validation of immunohistochemical assays: guideline from the College of American Pathologists Pathology and Laboratory Quality Center. Arch Pathol Lab Med. 2014;138(11):1432-1443. doi:10.5858/arpa.2013-0610-CP.
45.
FungKMMessingALeeVM-YTrojanowskiJQ. A novel modification of the avidin-biotin complex method for immunohistochemical studies of transgenic mice with murine monoclonal antibodies. J Histochem Cytochem. 1992;40(9):1319-1328. doi:10.1177/40.9.1506669.
46.
GabrielVZdyrskiCSahooDK, et al. Adult animal stem cell-derived organoids in biomedical research and the One Health paradigm. Int J Mol Sci. 2024;25(2):701. doi:10.3390/ijms25020701.
47.
GanJBolonBVan VleetTWoodC. Alternative models in biomedical research: in silico, in vitro, ex vivo, and non-traditional in vivo approaches. In: HaschekWMRousseauxCGWalligMABolonB, eds. Haschek and Rousseaux’s Handbook of Toxicologic Pathology. 4th ed. Vol. 1.San Diego, CA: Academic Press (Elsevier); 2022:925-966.
48.
GoldsmithJDFitzgibbonsPLSwansonPE. Principles of analytic validation of clinical immunohistochemistry assays. Adv Anat Pathol. 2015;22(6):384-387. doi:10.1097/PAP.0000000000000094.
49.
GoldsmithJDTroxellMLRoy-ChowdhuriS, et al. Principles of analytic validation of immunohistochemical assays: guideline update. Arch Pathol Lab Med. 2024;148(6):e111-e153. doi:10.5858/arpa.2023-0483-CP.
50.
GoldsteinNSHewittSMTaylorCRYazijiHHicksDG. Recommendations for improved standardization of immunohistochemistry. Appl Immunohistochem Mol Morphol. 2007;15(2):124-133. doi:10.1097/PAI.0b013e31804c7283.
51.
GrilloFBruzzoneMPigozziS, et al. Immunohistochemistry on old archival paraffin blocks: is there an expiry date?J Clin Pathol. 2017;70(11):988-993. doi:10.1136/jclinpath-2017-204387.
52.
GuerinCJ. Using antibodies in microscopy: a guide to immunohistochemistry. Part 2: IHC staining protocols. Microsc Today. 2023;31(3):34-39. doi:10.1093/mictod/qaad029.
53.
GündischSGrundner-CulemannKWolffC, et al. Delayed times to tissue fixation result in unpredictable global phosphoproteome changes. J Proteome Res. 2013;12(10):4424-4434. doi:10.1021/pr400451z.
HayatMA. Antigens and antibodies. In: Microscopy, Immunohistochemistry, and Antigen Retrieval Methods for Light and Electron Microscopy. Cham: Springer; 2002:31-55.
56.
HelanderKG. Formaldehyde binding in brain and kidney: a kinetic study of fixation. J Histotechnol. 1999;22(4):317-318. doi:10.1179/his.1999.22.4.317.
57.
HelanderKG. Kinetic studies of formaldehyde binding in tissue. Biotechnic Histochem. 1994;69(3):177-179. doi:10.3109/10520299409106282.
58.
HewittSMBaskinDGFrevertCWStahlWLRosa-MolinarE. Controls for immunohistochemistry: the Histochemical Society’s standards of practice for validation of immunohistochemical assays. J Histochem Cytochem. 2014;62(10):693-697. doi:10.1369/0022155414545224.
59.
HewlettB. Penetration rates of formaldehyde. Microsc Today. 2002;10(6):30-31. doi:10.1017/S1551929500058491.
60.
HinzNJuckerM. Distinct functions of AKT isoforms in breast cancer: a comprehensive review. Cell Commun Signal. 2019;17(1):154. doi:10.1186/s12964-019-0450-3.
61.
HolmsethSZhouYFollin-ArbeletVVLehreKPBerglesDEDanboltNC. Specificity controls for immunocytochemistry: the antigen preadsorption test can lead to inaccurate assessment of antibody specificity. J Histochem Cytochem. 2012;60(3):174-187. doi:10.1369/0022155411434828.
JanardhanKSJensenHClaytonNPHerbertRA. Immunohistochemistry in investigative and toxicologic pathology. Toxicol Pathol. 2018;46(5):488-510. doi:10.1177/0192623318776907.
65.
JohnsonCW. Issues in immunohistochemistry. Toxicol Pathol. 1999;27(2):246-248. doi:10.1177/019262339902700214.
66.
KaltenbachH-M. Statistical Design and Analysis of Biological Experiments. Cham: Springer; 2021.
67.
KapalczynskaMKolendaTPrzybylaW, et al. 2D and 3D cell cultures–a comparison of different types of cancer cell cultures. Arch Med Sci. 2018;14(4):910-919. doi:10.5114/aoms.2016.63743.
68.
KaptchukTJ. Effect of interpretive bias on research evidence. BMJ. 2003;326(7404):1453-1455. doi:10.1136/bmj.326.7404.1453.
69.
KawaiTIkegawaMOriDAkiraS. Decoding Toll-like receptors: recent insights and perspectives in innate immunity. Immunity. 2024;57(4):649-673. doi:10.1016/j.immuni.2024.03.004.
70.
KiernanJA. Formaldehyde, formalin, paraformaldehyde and glutaraldehyde: what they are and what they do. Microsc Today. 2000;8:8-13.
71.
KilkennyCBrowneWJCuthillICEmersonMAltmanDG. Improving bioscience research reporting: the ARRIVE guidelines for reporting animal research. PLoS Biol. 2010;8(6):e1000412. doi:10.1371/journal.pbio.1000412.
72.
KimSWRohJParkCS. Immunohistochemistry for pathologists: protocols, pitfalls, and tips. J Pathol Transl Med. 2016;50(6):411-418. doi:10.4132/jptm.2016.08.08.
73.
KohlerGMilsteinC. Derivation of specific antibody-producing tissue culture and tumor lines by cell fusion. Eur J Immunol. 1976;6(7):511-519. doi:10.1002/eji.1830060713.
74.
KomuraDIshikawaS. Machine learning methods for histopathological image analysis. Comput Struct Biotechnol J. 2018;16:34-42. doi:10.1016/j.csbj.2018.01.001.
75.
KreitzCFurleyPMemmertDSimonsDJ. Inattentional blindness and individual differences in cognitive abilities. PLoS ONE. 2015;10(8):e0134675. doi:10.1371/journal.pone.0134675.
LatendresseJRWarbrittionARJonassenHCreasyDM. Fixation of testes and eyes using a modified Davidson’s fluid: comparison with Bouin’s fluid and conventional Davidson’s fluid. Toxicol Pathol. 2002;30(4):524-533. doi:10.1080/01926230290105721.
78.
LaytonCBancroftJDSuvarnaSK. Fixation of tissues. In: SuvarnaSKLaytonCBancroftJD, eds. Bancroft’s Theory and Practice of Histological Techniques. 8th ed.Elsevier; 2019:40-63.
79.
LeachMWHalpernWGJohnsonCW, et al. Use of tissue cross-reactivity studies in the development of antibody-based biopharmaceuticals: history, experience, methodology, and future directions. Toxicol Pathol. 2010;38(7):1138-1166. doi:10.1177/0192623310382559.
80.
LevyJJSalasLAChristensenBCSriharanAVaickusLJ. PathFlowAI: a high-throughput workflow for preprocessing, deep learning and interpretation in digital pathology. Pac Symp Biocomput. 2020;25:403-414. doi:10.1142/9789811215636_0036.
81.
LinFPrichardJWLiuHWilkersonML. Handbook of Practical Immunohistochemistry. 3rd ed.Cham: Springer; 2022.
82.
MacLachlanTKPriceSCavagnaroJ, et al. Classic and evolving approaches to evaluating cross reactivity of mAb and mAb-like molecules–a survey of industry 2008-2019. Regul Toxicol Pharmacol. 2021;121:104872. doi:10.1016/j.yrtph.2021.104872.
83.
MacNeilTVathiotisIAMartinez-MorillaS, et al. Antibody validation for protein expression on tissue slides: a protocol for immunohistochemistry. Biotechniques. 2020;69(6):460-468. doi:10.2144/btn-2020-0095.
84.
MagakiSHojatSAWeiBSoAYongWH. An introduction to the performance of immunohistochemistry. Methods Mol Biol. 2019;1897:289-298. doi:10.1007/978-1-4939-8935-5_25.
85.
MarinopoulosAEAyresSCBiswasS, et al. Optimization of decalcification techniques for histologic examination of the rat maxillary and mandibular incisors for toxicity studies. J Histotechnol. 2022;45(1):2-9. doi:10.1080/01478885.2021.1974780.
86.
MetindirJDilekGBPakI. Staining characterization by immunohistochemistry of tumor cancer antigen in patients with endometrial cancer. Eur J Gynaecol Oncol. 2008;29(5):489-492.
87.
MeyerholzDKBeckAP. Fundamental concepts for semiquantitative tissue scoring in translational research. ILAR J. 2018;59(1):13-17. doi:10.1093/ilar/ily025.
88.
MeyerholzDKBeckAP. Principles and approaches for reproducible scoring of tissue stains in research. Lab Invest. 2018;98(7):844-855. doi:10.1038/s41374-018-0057-0.
89.
MeyerholzDKLambertzAMReznikovLR, et al. Immunohistochemical detection of markers for translational studies of lung disease in pigs and humans. Toxicol Pathol. 2016;44(3):434-441. doi:10.1177/0192623315609691.
90.
MeyerholzDKSierenJCBeckAPFlahertyHA. Approaches to evaluate lung inflammation in translational research. Vet Pathol. 2018;55(1):42-52. doi:10.1177/0300985817726117.
91.
MeyerholzDKStoltzDAGansemerND, et al. Lack of cystic fibrosis transmembrane conductance regulator disrupts fetal airway development in pigs. Lab Invest. 2018;98(6):825-838. doi:10.1038/s41374-018-0026-7.
92.
MeyerholzDKTintleNLBeckAP. Common pitfalls in analysis of tissue scores. Vet Pathol. 2019;56(1):39-42. doi:10.1177/0300985818794250.
93.
MighellAJHumeWJRobinsonPA. An overview of the complexities and subtleties of immunohistochemistry. Oral Dis. 1998;4(3):217-223. doi:10.1111/j.1601-0825.1998.tb00282.x.
94.
MooreHMKellyABJewellSD, et al. Biospecimen reporting for improved study quality (BRISQ). J Proteome Res. 2011;10(8):3429-3438. doi:10.1021/pr200021n.
95.
Morrison-BogoradMZimmermanALPardueS. Heat-shock 70 messenger RNA levels in human brain: correlation with agonal fever. J Neurochem. 1995;64(1):235-246. doi:10.1046/j.1471-4159.1995.64010235.x.
96.
NasoJRPovshednaTWangG, et al. Automated PD-L1 scoring for non-small cell lung carcinoma using open-source software. Pathol Oncol Res. 2021;27:609717. doi:10.3389/pore.2021.609717.
97.
NeefNNikulaKJFrancke-CarrollSBooneL. Regulatory Forum opinion piece: blind reading of histopathology slides in general toxicology studies. Toxicol Pathol. 2012;40(4):697-699. doi:10.1177/0192623312438737.
O’HurleyGSjöstedtERahmanA, et al. Garbage in, garbage out: a critical evaluation of strategies used for validation of immunohistochemical biomarkers. Mol Oncol. 2014;8(4):783-798. doi:10.1016/j.molonc.2014.03.008.
100.
PaavilainenLEdvinssonAAsplundA, et al. The impact of tissue fixatives on morphology and antibody-based protein profiling in tissues and cells. J Histochem Cytochem. 2010;58(3):237-246. doi:10.1369/jhc.2009.954321.
101.
Pinheiro da SilvaFGonçalvesANADuarte-NetoAN, et al. Transcriptome analysis of six tissues obtained post-mortem from sepsis patients. J Cell Mol Med. 2023;27(20):3157-3167. doi:10.1111/jcmm.17938.
102.
PottsSJKruegerJSLandisND, et al. Evaluating tumor heterogeneity in immunohistochemistry-stained breast cancer tissue. Lab Invest. 2012;92(9):1342-1357. doi:10.1038/labinvest.2012.91.
103.
PrasseKHildebrandtPDoddD, et al. Should the microscopic evaluation of slides from toxicity and carcinogenicity studies in animals be conducted in a “blind” fashion?Vet Pathol. 1986;23(4):540-541. doi:10.1177/030098588602300434.
104.
PriceSAMcDormanKChanC, et al. Special techniques in toxicologic pathology. In: Haschek and Rousseaux’s Handbook of Toxicologic Pathology, HaschekRousseaux (eds). Academic Press, 2022, pp. 335-393.
105.
Ramos-VaraJA. Technical aspects of immunohistochemistry. Vet Pathol. 2005;42(4):405-426. doi:10.1354/vp.42-4-405.
106.
Ramos-VaraJAKiupelMBaszlerT, et al. Suggested guidelines for immunohistochemical techniques in veterinary diagnostic laboratories. J Vet Diagn Invest. 2008;20(4):393-413. doi:10.1177/104063870802000401.
107.
Ramos-VaraJAMillerMA. When tissue antigens and antibodies get along: revisiting the technical aspects of immunohistochemistry—the red, brown, and blue technique. Vet Pathol. 2014;51(1):42-87. doi:10.1177/0300985813505879.
108.
Ramos-VaraJAWebsterJDDuSoldDMillerMA. Immunohistochemical evaluation of the effects of paraffin section storage on biomarker stability. Vet Pathol. 2014;51(1):102-109. doi:10.1177/0300985813476067.
109.
RizzardiAEJohnsonATVogelRI, et al. Quantitative comparison of immunohistochemical staining measured by digital image analysis versus pathologist visual scoring. Diagn Pathol. 2012;7:42. doi:10.1186/1746-1596-7-42.
110.
SaposnikGRedelmeierDRuffCCToblerPN. Cognitive biases associated with medical decisions: a systematic review. BMC Med Inform Decis Mak. 2016;16(1):138. doi:10.1186/s12911-016-0377-1.
111.
SaviFMBrierlyGIBaldwinJTheodoropoulosCWoodruffMA. Comparison of different decalcification methods using rat mandibles as a model. J Histochem Cytochem. 2017;65(12):705-722. doi:10.1369/0022155417733708.
112.
SchaferKAEighmyJFikesJD, et al. Use of severity grades to characterize histopathologic changes. Toxicol Pathol. 2018;46(3):256-265. doi:10.1177/0192623318761348.
113.
ScudamoreCLSoilleuxEJKarpNA, et al. Recommendations for minimum information for publication of experimental pathology data: MINPEPA guidelines. J Pathol. 2016;238(2):359-367. doi:10.1002/path.4642.
114.
SfanosKSYegnasubramanianSNelsonWG, et al. If this is true, what does it imply? How end-user antibody validation facilitates insights into biology and disease. Asian J Urol. 2019;6(1):10-25. doi:10.1016/j.ajur.2018.11.006.
115.
SkalandIOvestadIJanssenEAM, et al. Digital image analysis improves the quality of subjective HER-2 expression scoring in breast cancer. Appl Immunohistochem Mol Morphol. 2008;16(2):185-190. doi:10.1097/PAI.0b013e318059c20c.
116.
SnyderJMRadaelliEGoekenA, et al. Perfusion with 10% neutral-buffered formalin is equivalent to 4% paraformaldehyde for histopathology and immunohistochemistry in a mouse model of experimental autoimmune encephalomyelitis. Vet Pathol. 2022;59(3):498-505. doi:10.1177/03009858221075588.
117.
SouthgateJHarndenPTrejdosiewiczLK. Cytokeratin expression patterns in normal and malignant urothelium: a review of the biological and diagnostic implications. Histol Histopathol. 1999;14(2):657-664. doi:10.14670/HH-14.657.
118.
TaubeJMAkturkGAngeloM, et al. The Society for Immunotherapy of Cancer statement on best practices for multiplex immunohistochemistry (IHC) and immunofluorescence (IF) staining and validation. J Immunother Cancer. 2020;8(1):e000155. doi:10.1136/jitc-2019-000155.
119.
TaylorCR. Quantifiable internal reference standards for immunohistochemistry: the measurement of quantity by weight. Appl Immunohistochem Mol Morphol. 2006;14(3):253-259. doi:10.1097/00129039-200609000-00001.
120.
TaylorCR. Report of the Immunohistochemistry Steering Committee of the Biological Stain Commission. “Proposed format: package insert for immunohistochemistry products.”Biotechnic Histochem. 1992;67(6):323-338. doi:10.3109/10520299209110045.
121.
TaylorCRLevensonRM. Quantification of immunohistochemistry—issues concerning methods, utility and semiquantitative assessment II. Histopathology. 2006;49(4):411-424. doi:10.1111/j.1365-2559.2006.02513.x.
TaylorCRShiS-R. Techniques of immunohistochemistry: principles, pitfalls, and standardization. In: DabbsDJ, ed. Diagnostic Immunohistochemistry: Theranostic and Genomic Applications. 4th ed.Elsevier (Saunders); 2014:1-41.
124.
TomitaHVawterMPWalshDM, et al. Effect of agonal and postmortem factors on gene expression profile: quality control in microarray analyses of postmortem human brain. Biol Psychiatr. 2004;55(4):346-352. doi:10.1016/j.biopsych.2003.10.013.
125.
TorlakovicEENielsenSVybergMTaylorCR. Getting controls under control: the time is now for immunohistochemistry. J Clin Pathol. 2015;68(11):879-882. doi:10.1136/jclinpath-2014-202705.
126.
TorlakovicEERiddellRBanerjeeD, et al. Canadian Association of Pathologists–Association canadienne des pathologistes National Standards Committee/Immunohistochemistry: best practice recommendations for standardization of immunohistochemistry tests. Am J Clin Pathol. 2010;133(3):354-365. doi:10.1309/AJCPDYZ1XMF4HJWK.
127.
UhlenMBandrowskiACarrS, et al. A proposal for validation of antibodies. Nat Methods. 2016;13(10):823-827. doi:10.1038/nmeth.3995.
128.
US Code of Federal Regulations (CFR). Title 50-Wildlife and Fisheries. Chapter I - United States Fish and Wildlife Service, Department of the Interior. Subchapter B - Taking, possession, transportation, sale, purchase, barter, exportation, and importation of wildlife and plants. Part 23 - Convention on International Trade in Endangered Species of wild fauna and flora (CITES). US Government Publishing Office. Accessed November 30, 2025. https://www.ecfr.gov/current/title-50/chapter-I/subchapter-B/part-23.
van der LoosCM. Multiple immunoenzyme staining: methods and visualizations for the observation with spectral imaging. J Histochem Cytochem. 2008;56(4):313-328. doi:10.1369/jhc.2007.950170.
132.
WalkerRA. Quantification of immunohistochemistry—issues concerning methods, utility and semiquantitative assessment I. Histopathology. 2006;49(4):406-410. doi:10.1111/j.1365-2559.2006.02514.x.
133.
WebsterJDSolonMGibson-CorleyKN. Validating immunohistochemistry assay specificity in investigative studies: considerations for a weight of evidence approach. Vet Pathol. 2021;58(5):829-840. doi:10.1177/0300985820960132.
134.
XieRChungJYYlayaK, et al. Factors influencing the degradation of archival formalin-fixed paraffin-embedded tissue sections. J Histochem Cytochem. 2011;59(4):356-365. doi:10.1369/0022155411398488.
