Medicinal plants are still being used as therapeutic agents in many developing nations because they are more widely available. This research aimed to compare method /result reliability between poison plate and agar well techniques in determining antifungal activity of protein fractions of plant samples. The phytochemicals constituents present in the leaves (Moringa olifera, Vernonia amygdalina, Annona muricata and seeds/roots of Garcinia kola, Buccholzia coricea, Allium sativum and Zingiber officinale) was extracted by air drying and subjecting them to maceration in 75% ammonium sulphate. Thereafter, a sensitivity test was carried out using two methods; poisoned food technique to determine antifungal activity and agar well diffusion. The in vitro antimicrobial susceptibility test of the protein fractions was treated against selected fungi isolates (Penicilium, Aspargilus and Rhizopus). The results showed that the antifungal activities of Garcinia kola against selected microbes, at 100 mg for Aspergillus 1, Penicillin 1, Penicillin 3, Rhizopus and Aspergillus 3 for agar well showed, 0.00 ± 0.00, 18.1 ± 0.16, 18.0 ± 0.16, 20.0 ± 0.50, 0.00 ± 0.00, while in poison plate they exhibited 16.0 ± 0.00, 20.1 ± 0.16, 19.8 ± 0.28, 20.0 ± 2.00, 16.0 ± 0.50 respectively for each method. Also, the antifungal activities of Zingiber officinale showed a significant difference (p < 0.05) in poison plate technique than agar well when the concentration of the microbes where 100 mg, except for Rhizopus which showed similar result for both poison plate and agar well technique (15.8 + 0.16). The findings of the current research showed that poison plate technique is more sensitive to the antifungal activities in the protein fractions when compared to the agar well diffusion.